Substituted carbamoylmethylamino acetic acid derivatives as novel NEP inhibitors

ABSTRACT

The present invention provides a compound of formula I; 
     
       
         
         
             
             
         
       
     
     or a pharmaceutically acceptable salt thereof, wherein R 1 , R 2 , R 3 , R 4 , R 6 , A 1 , A 2 , X 1 , s and m are defined herein. The invention also relates to a method for manufacturing the compounds of the invention, and its therapeutic uses. The present invention further provides a combination of pharmacologically active agents and a pharmaceutical composition.

BACKGROUND OF THE INVENTION

Endogenous atrial natriuretic peptides (ANP), also called atrialnatriuretic factors (ANF) have diuretic, natriuretic and vasorelaxantfunctions in mammals. The natural ANF peptides is metabolicallyinactivated, in particular by a degrading enzyme which has beenrecognized to correspond to the enzyme neutral endopeptidase (NEP) EC3.4.24.11, also responsible for e.g. the metabolic inactivation ofenkephalins.

Neutral endopeptidase (EC 3.4.24.11; enkephalinase; atriopeptidase; NEP)is a zinc-containing metalloprotease that cleaves a variety of peptidesubstrates on the amino side of hydrophobic residues [see Pharmacol Rev,Vol. 45, p. 87 (1993)]. Substrates for this enzyme include, but are notlimited to, atrial natriuretic peptide (ANP, also known as ANF), brainnatriuretic peptide (BNP), met- and leu-enkephalin, bradykinin,neurokinin A, endothelin-1 and substance P. ANP is a potent vasorelaxantand natriuretic agent [see J Hypertens, Vol. 19, p. 1923 (2001)].Infusion of ANP in normal subjects resulted in a reproducible, markedenhancement of natriuresis and diuresis, including increases infractional excretion of sodium, urinary flow rate and glomerularfiltration rate [see J Clin Pharmacol, Vol. 27, p. 927 (1987)]. However,ANP has a short half-life in circulation, and NEP in kidney cortexmembranes has been shown to be the major enzyme responsible fordegrading this peptide [see Peptides, Vol. 9, p. 173 (1988)]. Thus,inhibitors of NEP (neutral endopeptidase inhibitors, NEPi) shouldincrease plasma levels of ANP and, hence, are expected to inducenatriuretic and diuretic effects.

This enzyme is involved in the breakdown of several bioactiveoligopeptides, cleaving peptide bonds on the amino side of hydrophobicamino acid residues. The peptides metabolised include atrial natriureticpeptides (ANP), bombesin, bradykinin, calcitonin gene-related peptide,endothelins, enkephalins, neurotensin, substance P and vasoactiveintestinal peptide. Some of these peptides have potent vasodilatory andneurohormone functions, diuretic and natriuretic activity or mediatebehaviour effects.

SUMMARY OF THE INVENTION

The aim of the present invention is to provide novel compounds which areuseful as neutral endopeptidase inhibitors, e.g. as inhibitors of theANF-degrading enzyme in mammals, so as to prolong and potentiate thediuretic, natriuretic and vasodilator properties of ANF in mammals, byinhibiting the degradation thereof to less active metabolites.

The compounds of this invention are thus particularly useful for thetreatment of conditions and disorders responsive to the inhibition ofneutral endopeptidase (NEP) EC 3.4.24.11.

The invention pertains to the compounds, methods for using them, anduses thereof as described herein. Examples of compounds of the inventioninclude the compounds according to anyone of Formulae I′ and I to IV, ora pharmaceutically acceptable salt thereof and the compounds of theexamples.

The invention therefore provides a compound of the formula (I′):

wherein:X¹ is OH, —O—C₁₋₇alkyl, —NR^(a)R^(b), —NHS(O)₂—C₁₋₇alkyl or—NHS(O)₂-benzyl, wherein R^(a) and R^(b) for each occurrence areindependently H or C₁₋₇alkyl;R¹ is H, C₁₋₆ alkyl or C₆₋₁₀-aryl-C₁₋₆ alkyl, wherein alkyl isoptionally substituted with benzyloxy, hydroxy or C₁₋₆ alkoxy; for eachoccurence, R² is independently C₁₋₆-alkoxy, hydroxy, halo, C₁₋₆-alkyl,cyano or trifluoromethyl;

A² is O or NR⁵;

R⁴ and R⁶ are independently H or C₁₋₆ alkyl;A¹ is a bond or C₁₋₃alkylene chain;R³ is a 5- or 6-membered heteroaryl, C₆₋₁₀-aryl or C₃₋₇-cycloalkyl,wherein each heteroaryl, aryl or cycloalkyl are optionally substitutedwith one or more groups independently selected from the group consistingof C₁₋₆alkyl, halo, haloC₁₋₆alkyl, C₁₋₆alkoxy, hydroxy, CO₂H andCO₂C₁₋₆alkyl;R⁶ for each occurrence is independently halo, hydroxy, C₁₋₇alkoxy, halo,C₁₋₇alkyl or halo-C₁₋₇alkyl; orR⁴, A¹-R³, together with the nitrogen to which R⁴ and A¹-R³ areattached, form a 4- to 7-membered heterocyclyl or a 5- to 6-memberedheteroaryl, each of which is optionally substituted with one or moregroups independently selected from the group consisting of C₁₋₆alkyl,halo, haloC₁₋₆alkyl, C₁₋₆alkoxy, hydroxy, CO₂H and CO₂C₁₋₆alkyl; andm is 0 or an integer from 1 to 5;s is 0 or an integer from 1 to 4; ora pharmaceutically acceptable salt thereof.

The invention also provides a compound of the formula (I):

wherein:X¹ represent OH or O—C₁₋₆-alkyl;R¹ is H, C₁₋₆ alkyl or C₆₋₁₀-aryl-C₁₋₆ alkyl; for each occurence, R² isindependently C₁₋₆-alkoxy, hydroxy, halo, C₁₋₆-alkyl, cyano ortrifluoromethyl;R⁴ and R⁵ are independently H or C₁₋₆ alkyl;A¹ is a bond or C₁₋₃alkylene chain;R³ is a 5- or 6-membered heteroaryl, C₆₋₁₀-aryl or C₃₋₇-cycloalkyl,wherein each heteroaryl, aryl or cycloalkyl are optionally substitutedwith one or more groups independently selected from the group consistingof C₁₋₆alkyl, halo, haloC₁₋₆alkyl, C₁₋₆alkoxy, hydroxy, CO₂H andCO₂C₁₋₆alkyl;R⁶ for each occurrence is independently halo, hydroxy, C₁₋₇alkoxy, halo,C₁₋₇alkyl or halo-C₁₋₇alkyl; orR⁴, A¹-R³, together with the nitrogen to which R⁴ and A¹-R³ areattached, form a 4- to 7-membered heterocyclyl or a 5- to 6-memberedheteroaryl, each of which is optionally substituted with one or moregroups independently selected from the group consisting of C₁₋₈alkyl,halo, haloC₁₋₆alkyl, C₁₋₆alkoxy, hydroxy, CO₂H and CO₂C₁₋₆alkyl; andm is 0 or an integer from 1 to 5;s is 0 or an integer from 1 to 4; ora pharmaceutically acceptable salt thereof.

The compounds of the invention, by inhibiting the neutral endopeptidaseEC.3.4.24.11, can potentiate the biological effects of bioactivepeptides. Thus, in particular the compounds have utility in thetreatment of a number of disorders, including hypertension, resistanthypertension, pulmonary hypertension, pulmonary arterial hypertension,isolated systolic hypertension, peripheral vascular disease, heartfailure, congestive heart failure, left ventricular hypertrophy, angina,renal insufficiency (diabetic or non-diabetic), renal failure (includingedema and salt retension), diabetic nephropathy, non-diabeticnephropathy, nephroic syndrome, glomerulonephritis, scleroderma,glomerular sclerosis, proteinurea of primary renal disease, renalvascular hypertention, diabetic retinopathy and end-stage renal disease(ESRD), endothelial dysfunction, diastolic dysfunction, hypertrophiccardiomyopathy, diabetic cardiac myopathy, supraventricular andventricular arrhythmias, atrial fibrillation (AF), cardiac fibrosis,atrial flutter, detrimental vascular remodeling, plaque stabilization,myocardial infarction (MI), renal fibrosis, polycystic kidney disease(PKD), renal failure (including edema and salt retension), cyclicaloedema, Menières disease, hyperaldosteroneism (primary and secondary)and hypercalciuria, ascites. In addition, because of their ability topotentiate the effects of ANF the compounds have utility in thetreatment of glaucoma. As a further result of their ability to inhibitthe neutral endopeptidase E.C.3.4.24.11 the compounds of the inventionmay have activity in other therapeutic areas including for example thetreatment of menstrual disorders, preterm labour, pre-eclampsia,endometriosis, and reproductive disorders (especially male and femaleinfertility, polycystic ovarian syndrome, implantation failure). Alsothe compounds of the invention should treat asthma, obstructive sleepapnea, inflammation, leukemia, pain, epilepsy, affective disorders suchas depression and psychotic condition such as dementia and geriatricconfusion, obesity and gastrointestinal disorders (especially diarrheaand irritable bowel syndrome), wound healing (especially diabetic andvenous ulcers and pressure sores), septic shock, gastric acid secretiondysfunction, hyperreninaemia, cystic fibrosis, restenosis, type-2diabetes, metabolic syndrome, diabetic complications andathereosclerosis, male and female sexual dysfunction.

In a preferred embodiment the compounds of the invention are useful inthe treatment of cardiovascular disorders.

In another embodiment, the invention pertains to a method for treating adisorders or diseases responsive to the inhibition of neutralendopeptidase EC 3.4. 24.11 (NEP), in a subject in need of suchtreatment, comprising: administering to the subject an effective amountof a compound according to anyone of Formulae I-IV, or apharmaceutically acceptable salt thereof, such that the disorder ordisease responsive to the inhibition of neutral endopeptidase EC 3.4.24.11 (NEP) in the subject is treated.

In yet another embodiment, the invention pertains to pharmaceuticalcompositions, comprising a compound according to anyone of FormulaeI-IV, or a pharmaceutically acceptable salt thereof, and one or morepharmaceutically acceptable carriers.

In still another embodiment, the invention pertains to combinationsincluding, a compound according to anyone of Formulae I-IV, or apharmaceutically acceptable salt thereof, and pharmaceuticalcombinations of one or more therapeutically active agents.

In another embodiment, the invention pertains to a method for inhibitingneutral endopeptidase EC 3.4. 24.11 in a subject in need thereof,comprising: administering to the subject a therapeutically effectiveamount of a compound according to anyone of Formulae I-IV, or apharmaceutically acceptable salt thereof, such that neutralendopeptidase EC 3.4. 24.11 is inhibited.

DETAILED DESCRIPTION OF THE INVENTION Definition

For purposes of interpreting this specification, the followingdefinitions will apply unless specified otherwise and wheneverappropriate, terms used in the singular will also include the plural andvice versa.

As used herein, the term “alkyl” refers to a fully saturated branched orunbranched (or straight chain or linear) hydrocarbon moiety, comprising1 to 20 carbon atoms. Preferably the alkyl comprises 1 to 6 carbonatoms, and more preferably 1 to 4 carbon atoms. Representative examplesof alkyl include methyl, ethyl, n-propyl, iso-propyl, n-butyl,sec-butyl, iso-butyl, tert-butyl, n-pentyl, isopentyl, neopentyl,n-hexyl, 3-methylhexyl, 2,2-dimethylpentyl, 2,3-dimethylpentyl,n-heptyl. The term “C₁₋₆alkyl” refers to a hydrocarbon having from oneto six carbon atoms. The term “alkylene” refers to a divalent alkylradical, wherein alkyl is as previously defined.

As used herein, the term “haloalkyl” refers to an alkyl as definedherein, that is substituted by one or more halo groups as definedherein. Preferably the haloalkyl can be monohaloalkyl, dihaloalkyl orpolyhaloalkyl including perhaloalkyl. A monohaloalkyl can have one iodo,bromo, chloro or fluoro within the alkyl group. Dihaloalky andpolyhaloalkyl groups can have two or more of the same halo atoms or acombination of different halo groups within the alkyl. Preferably, thepolyhaloalkyl contains up to 12, or 10, or 8, or 6, or 4, or 3, or 2halo groups. Representative examples of haloalkyl are fluoromethyl,difluoromethyl, trifluoromethyl, chloromethyl, dichloromethyl,trichloromethyl, pentafluoroethyl, heptafluoropropyl,difluorochloromethyl, dichlorofluoromethyl, difluoroethyl,difluoropropyl, dichloroethyl and dichloropropyl. A perhaloalkyl refersto an alkyl having all hydrogen atoms replaced with halo atoms. The term“halo-C₁₋₆alkyl” refers to a hydrocarbon having one to six carbon atomsand being substituted by one or more halo groups.

As used herein, the term “alkoxy” refers to alkyl-O—, wherein alkyl isdefined herein above. Representative examples of alkoxy include, but arenot limited to, methoxy, ethoxy, propoxy, 2-propoxy, butoxy,tert-butoxy, pentyloxy, hexyloxy, cyclopropyloxy-, cyclohexyloxy- andthe like. Preferably, alkoxy groups have about 1-6, more preferablyabout 1-4 carbons.

As used herein, the term “cycloalkyl” refers to saturated or partiallyunsaturated monocyclic, bicyclic or tricyclic hydrocarbon groups of 3-12carbon atoms, preferably 3-8, or 3-7 carbon atoms. For bicyclic, andtricyclic cycloalkyl system, all rings are non-aromatic. Exemplarymonocyclic hydrocarbon groups include cyclopropyl, cyclobutyl,cyclopentyl, cyclopentenyl, cyclohexyl and cyclohexenyl. Exemplarybicyclic hydrocarbon groups include bornyl, decahydronaphthyl,bicyclo[2.1.1]hexyl, bicyclo[2.2.1]heptyl, bicyclo[2.2.1]heptenyl,bicyclo[2.2.2]octyl. Exemplary tricyclic hydrocarbon groups includeadamantyl. The term “C₃₋₇cycloakyl” refers to a cyclic hydrocarbongroups having 3 to 7 carbon atoms.

The term “aryl” refers to monocyclic or bicyclic aromatic hydrocarbongroups having 6-10 carbon atoms in the ring portion. The term “aryl”also refer to a group in which the aromatic ring is fused to acycloalkyl ring, where the radical of attachment is on the aromatic ringor on the fused cycloalkyl ring. Representative examples of aryl arephenyl, naphthyl, hexahydroindyl, indanyl or tetrahydronaphthyl. Theterm “C₆₋₁₀aryl” refers to an aromatic hydrocarbon groups having 6 to 10carbon atoms in the ring portion.

The term “arylalkyl” is an alkyl substituted with aryl. Representativeexamples of arylalkyl are benzyl or Phenyl-CH₂CH₂—. The term“C₆₋₁₀aryl-C₁₋₆alkyl” refers to a hydrocarbon having one to six carbonatoms, which hydrocarbon is substituted with an aryl having 6 to 10carbon atoms.

The term “Heteroaryl” includes monocyclic or bicyclic heteroaryl,containing from 5-10 ring members selected from carbon atoms and 1 to 5heteroatoms, and each heteroatoms is independently selected from O, N orS wherein S and N may be oxidized to various oxidation states. Forbicyclic heteroaryl system, the system is fully aromatic (i.e. all ringsare aromatic).

Typical monocyclic heteroaryl groups include thienyl, furyl, pyrrolyl,imidazolyl, pyrazolyl, thiazolyl, isothiazolyl, 1,2,3-oxadiazolyl,1,2,4-oxadiazolyl, 1,2,5-oxadiazolyl, 1,3,4-oxadiazolyl,1,2,3-thiadiazolyl, 1,2,4-thiadiazolyl, 1,2,5-thiadiazolyl,1,3,4-thiadiazolyl, isothiazol-3-yl, isothiazol-4-yl, isothiazol-5-yl,oxazol-2-yl, oxazol-4-yl, oxazol-5-yl, isoxazol-3-yl, isoxazol-4-yl,isoxazol-5-yl, 1,2,4-triazol-3-yl, 1,2,4-triazol-5-yl,1,2,3-triazol-4-yl, 1,2,3-triazol-5-yl, tetrazolyl, pyrid-2-yl,pyrid-3-yl, or pyridyl-4-yl, pyridazin-3-yl, pyridazin-4-yl,pyrazin-3-yl, 2-pyrazin-2-yl, pyrazin-4-yl, pyrazin-5-yl, 2-, 4-, or5-pyrimidin-2-yl, pyrimidin-4-yl, pyrimidin-5-yl. The term “heteroaryl”also refers to a group in which a heteroaromatic ring is fused to one ormore aryl rings, where the radical or point of attachment is on theheteroaromatic ring or on the fused aryl ring. Representative examplesof bicyclic heteroaryl are indolyl, isoindolyl, indazolyl, indolizinyl,purinyl, quinolizinyl, quinolinyl, isoquinolinyl, cinnolinyl,phthalazinyl, naphthyridinyl, quinazolinyl, quinaxalinyl,phenanthridinyl, phenathrolinyl, phenazinyl, phenothiazinyl,phenoxazinyl, benzisoqinolinyl, thieno[2,3-b]furanyl,furo[3,2-b]-pyranyl, 5H-pyrido[2,3-d]-o-oxazinyl,1H-pyrazolo[4,3-d]-oxazolyl, 4H-imidazo[4,5-d]thiazolyl,pyrazino[2,3-d]pyridazinyl, imidazo[2,1-b]thiazolyl,imidazo[1,2-b][1,2,4]triazinyl, 7-benzo[b]thienyl, benzoxazolyl,benzimidazolyl, benzothiazolyl, benzoxapinyl, benzoxazinyl,1H-pyrrolo[1,2-b][2]benzazapinyl, benzofuryl, benzothiophenyl,benzotriazolyl, pyrrolo[2,3-b]pyridinyl, pyrrolo[3,2-c]pyridinyl,pyrrolo[3,2-c]pyridinyl, pyrrolo[3,2-b]pyridinyl,imidazo[4,5-b]pyridinyl, imidazo[4,5-c]pyridinyl,pyrazolo[4,3-d]pyridinyl, pyrazolo[4,3-c]pyridinyl,pyrazolo[3,4-c]pyridinyl, pyrazolo[3,4-d]pyridinyl,pyrazolo[3,4-b]pyridinyl, imidazo[1,2-a]pyridinyl,pyrazolo[1,5-a]pyridinyl, pyrrolo[1,2-b]pyridazinyl,imidazo[1,2-c]pyrimidinyl, pyrido[3,2-d]pyrimidinyl,pyrido[4,3-d]pyrimidinyl, pyrido[3,4-d]pyrimidinyl,pyrido[2,3-d]pyrimidinyl, pyrido[2,3-b]pyrazinyl,pyrido[3,4-b]pyrazinyl, pyrimido[5,4-d]pyrimidinyl,pyrazino[2,3-b]pyrazinyl, or pyrimido[4,5-d]pyrimidinyl. When aheteroaryl moiety is substituted with hydroxy, the invention alsopertains to its oxo tautomeric. For example, an oxadiazole substitutedwith hydroxy also includes oxo-oxadiazole also known as oxadiazolone.The tautomerisation is represented as follow:

As used herein, the term “heterocyclyl” or “heterocyclo” refers to anoptionally substituted, saturated or unsaturated non-aromatic (partiallyunsaturated) ring which is a 4-, 5-, 6-, or 7-membered monocyclic, andcontains at least one heteroatom selected from O, S and N, where the Nand S can also optionally be oxidized to various oxidation states. Forbicyclic and tricyclic heterocyclyl ring system, a non-aromatic ringsystem is defined as being a non-fully or partially unsaturated ringsystem. Therefore bicyclic and tricyclic heterocyclyl ring systemsincludes heterocyclyl ring systems wherein one of the fused rings isaromatic but the other(s) is (are) non-aromatic. In one embodiment,heterocyclyl moiety represents a saturated monocyclic ring containingfrom 5-7 ring atoms and optionally containing a further heteroatom,selected from O, S or N. The heterocyclic group can be attached at aheteroatom or a carbon atom. The heterocyclyl can include fused orbridged rings as well as spirocyclic rings. Examples of heterocyclesinclude dihydrofuranyl, dioxolanyl, dioxanyl, dithianyl, piperazinyl,pyrrolidine, dihydropyranyl, oxathiolanyl, dithiolane, oxathianyl,thiomorpholino, oxiranyl, aziridinyl, oxetanyl, oxepanyl, azetidinyl,tetrahydrofuranyl, tetrahydrothiophenyl, pyrrolidinyl,tetrahydropyranyl, piperidinyl, morpholino, piperazinyl, azepinyl,oxapinyl, oxaazepanyl, oxathianyl, thiepanyl, azepanyl, dioxepanyl, anddiazepanyl.

The term “halogen” includes fluorine, bromine, chlorine and iodine. Theterm “perhalogenated” generally refers to a moiety wherein all hydrogensare replaced by halogen atoms.

The term “heteroatom” includes atoms of any element other than carbon orhydrogen. Preferred heteroatoms are nitrogen, oxygen, sulfur andphosphorus. In one embodiment the heteroatoms is selected from N, O andS.

Compound of the Invention

Various embodiments of the invention are described herein. It will berecognized that features specified in each embodiment may be combinedwith other specified features to provide further embodiments.

Certain compounds of Formula I′ or I include compounds of Formula II:

or a pharmaceutically acceptable salt thereof, wherein R¹, R², R³, R⁴,R⁵, R⁶, X¹, A¹, s and m have the definition of Formula I, supra.

In one embodiment the invention pertains to compounds of formula I′, Ior II wherein:

X¹ represent OH or O—C₁₋₆-alkyl;R¹ is H or C₁₋₆ alkyl;for each occurrence, R² is independently C₁₋₆-alkoxy, hydroxy, halo,C₁₋₆-alkyl, cyano or trifluoromethyl;R⁴ and R⁵ are independently H or C₁₋₆ alkyl;A¹ is a bond or C₁₋₃alkylene chain;R³ is a 5- or 6-membered heteroaryl or C₆₋₁₀-aryl, wherein eachheteroaryl and aryl are optionally substituted with one or moresubstituents independently selected from the group consisting ofC₁₋₆alkyl, halo, haloC₁₋₆alkyl, C₁₋₆alkoxy, hydroxy, CO₂H andCO₂C₁₋₆alkyl;R⁶ for each occurrence is independently halo, hydroxy, C₁₋₇alkoxy, halo,C₁₋₇alkyl or halo-C₁₋₇alkyl;m is 0 or an integer from 1 to 5;s is 0 or an integer from 1 to 4; ora pharmaceutically acceptable salt thereof.

In another embodiment the invention pertains to compounds of formula I′,I or II

wherein:X¹ represent OH or O—C₁₋₆-alkyl;R¹ is H or C₁₋₆ alkyl;for each occurrence, R² is independently C₁₋₆-alkoxy, hydroxy, halo,C₁₋₆-alkyl, cyano or trifluoromethyl;R⁴ and R⁵ are independently H or C₁₋₆ alkyl;A¹ is a bond or C₁₋₃alkylene chain;R³ is a 5- or 6-membered heteroaryl optionally substituted with one ormore substituents independently selected from the group consisting ofC₁₋₆alkyl, halo, haloC₁₋₆alkyl, C₁₋₆alkoxy, hydroxy, CO₂H andCO₂C₁₋₆alkyl;

R⁶ for each occurrence is independently halo, hydroxy, C₁₋₇alkoxy, halo,C₁₋₇alkyl or halo-C₁₋₇alkyl;

m is 0 or an integer from 1 to 5;s is 0 or an integer from 1 to 4; ora pharmaceutically acceptable salt thereof.

Certain compounds of Formula I′ or I include compounds of Formula III:

or a pharmaceutically acceptable salt thereof, wherein R¹, R², R³, R⁴,R⁵, R⁶, X¹, A¹ and s have the definition of Formula I, supra and X² ishalo and p is 0 or an integer from 1 to 4.

Certain compounds of Formula I′, I, II or III include compounds ofFormula IV:

or a pharmaceutically acceptable salt thereof, wherein R¹, R², R³, R⁴,R⁵, R⁶, X¹, A¹, X², s and p have the definition of Formulae I, II andIII, supra.

In one embodiment, the invention pertains to compounds of Formula III orIV, or a pharmaceutically acceptable salt thereof, wherein X² is a Cl.In a further aspect of this embodiment p is 0.

The following embodiments can be used independently, collectively or inany combination or sub-combination:

In one embodiment, the invention pertains to compounds according toanyone of Formulae I′ and I to IV, or any of any other classes andsubclasses described supra, or a pharmaceutically acceptable saltthereof, wherein A¹ is a bond or CH₂. In a further embodiment A¹ is abond.

In another embodiment, the invention pertains to compounds according toanyone of Formulae I′ and I to IV, or any of any other classes andsubclasses described supra, or a pharmaceutically acceptable saltthereof, wherein R³ is an optionally substituted 5- or 6-memberedheteroaryl. In one aspect of this embodiment, R³ is a 6-membered ringheteroaryl selected from the group consisting of pyrazine, pyridine,pyrimidine, pyranone (e.g. optionally substituted pyran-4-one,pyran-2-one such as 3-hydroxy-pyran-4-one, 3-hydroxy-pyran-2-one),pyrimidinone and pyridinone (e.g. optionally substituted pyridin-4-oneor pyridin-2-one such as for example 3-hydroxy-1-methyl-pyridin-4-one or1-benzyl-pyridin-2-one). In another aspect of this embodiment R³ is a5-membered ring heteroaryl selected from the group consisting ofoxazole, pyrrole, pyrazole, isooxazole, triazole, tetrazole, oxadiazole(e.g. 1-oxa-3,4-diazole, 1-oxa-2,4-diazole), oxadiazolone (e.g.oxadiazol-2-one), thiazole, isothiazole, thiophene, imidazole andthiadiazole. Other representative examples of R³ are oxazolone,thiazolone, oxadiazolone triazolone, oxazolone, imidazolone, pyrazolone.In a further embodiment, the optional substituents on the heteroaryl areindependently C₁₋₆alkyl, halo, haloC₁₋₆alkyl, C₁₋₆alkoxy, hydroxy, CO₂Hor CO₂C₁₋₆alkyl.

In another aspect of the above embodiment, the invention pertains tocompounds according to anyone of Formulae I′ and I to IV, or any of anyother classes and subclasses described supra, or a pharmaceuticallyacceptable salt thereof, wherein R³ is tetrazole.

In another embodiment, the invention pertains to compounds according toanyone of Formulae I′ and I to IV, or any of any other classes andsubclasses described supra, or a pharmaceutically acceptable saltthereof, wherein R³ is an optionally substituted phenyl. In a furtherembodiment, the optional substituents on phenyl are independentlyC₁₋₆alkyl, halo, haloC₁₋₆alkyl, C₁₋₆alkoxy, hydroxy, CO₂H orCO₂C₁₋₆alkyl. In a further embodiment, phenyl is substituted with CO₂Hand further optionally substituted.

In another embodiment, the invention pertains to compounds according toanyone of Formulae I′ and I to IV or any of any other classes andsubclasses described supra, or a pharmaceutically acceptable saltthereof, wherein R¹ is C₁₋₆ alkyl (i.e. methyl, ethyl, propyl,isopropyl).

In another embodiment, the invention pertains to compounds according toanyone of Formulae I′ and I to IV or any of any other classes andsubclasses described supra, or a pharmaceutically acceptable saltthereof, wherein R⁴ is H.

In another embodiment, the invention pertains to compounds according toanyone of Formulae I′ and I to IV or any of any other classes andsubclasses described supra, or a pharmaceutically acceptable saltthereof, wherein R⁵ is H.

In another embodiment, the invention pertains to compounds according toanyone of Formulae I′ and I to IV or any of any other classes andsubclasses described supra, or a pharmaceutically acceptable saltthereof, wherein s is 0.

In another embodiment R¹, R², R³, R⁴, R⁵, R⁶, X¹, A¹, X², m, s and pgroups are those defined by the R¹, R², R³, R⁴, R⁵, R⁶, X¹, A¹, X², A¹,m, s and p groups in the Examples section below.

In another embodiment individual compounds according to the inventionare those listed in the Examples section below or a pharmaceuticallyacceptable salt thereof.

It will be noted that the structure of some of the compounds of thisinvention includes asymmetric carbon atoms. It is to be understoodaccordingly that the isomers arising from such asymmetry (e.g., allenantiomers and diastereomers) are included within the scope of thisinvention, unless indicated otherwise. Such isomers can be obtained insubstantially pure form by classical separation techniques and bystereochemically controlled synthesis. Furthermore, the structures andother compounds and moieties discussed in this application also includeall tautomers thereof.

As used herein, the term “isomers” refers to different compounds thathave the same molecular formula but differ in arrangement andconfiguration of the atoms. Also as used herein, the term “an opticalisomer” or “a stereoisomer” refers to any of the various stereo isomericconfigurations which may exist for a given compound of the presentinvention and includes geometric isomers. It is understood that asubstituent may be attached at a chiral center of a carbon atom.Therefore, the invention includes enantiomers, diastereomers orracemates of the compound. “Enantiomers” are a pair of stereoisomersthat are non-superimposable mirror images of each other. A 1:1 mixtureof a pair of enantiomers is a “racemic” mixture. The term is used todesignate a racemic mixture where appropriate. “Diastereoisomers” arestereoisomers that have at least two asymmetric atoms, but which are notmirror-images of each other. The absolute stereochemistry is specifiedaccording to the Cahn-IngoId-Prelog R-S system. When a compound is apure enantiomer the stereochemistry at each chiral carbon may bespecified by either R or S. Resolved compounds whose absoluteconfiguration is unknown can be designated (+) or (−) depending on thedirection (dextro- or levorotatory) which they rotate plane polarizedlight at the wavelength of the sodium D line. Certain of the compoundsdescribed herein contain one or more asymmetric centers or axes and maythus give rise to enantiomers, diastereomers, and other stereoisomericforms that may be defined, in terms of absolute stereochemistry, as (R)-or (S)-. The present invention is meant to include all such possibleisomers, including racemic mixtures, optically pure forms andintermediate mixtures. Optically active (R)- and (S)-isomers may beprepared using chiral synthons or chiral reagents, or resolved usingconventional techniques. If the compound contains a double bond, thesubstituent may be E or Z configuration. If the compound contains adisubstituted cycloalkyl, the cycloalkyl substituent may have a cis- ortrans-configuration. All tautomeric forms are also intended to beincluded.

Any asymmetric atom (e.g., carbon or the like) of the compound(s) of thepresent invention can be present in racemic or enantiomericallyenriched, for example the (R)-, (S)- or (R,S)-configuration. In certainembodiments, each asymmetric atom has at least 50% enantiomeric excess,at least 60% enantiomeric excess, at least 70% enantiomeric excess, atleast 80% enantiomeric excess, at least 90% enantiomeric excess, atleast 95% enantiomeric excess, or at least 99% enantiomeric excess inthe (R)- or (S)-configuration. Substituents at atoms with unsaturatedbonds may, if possible, be present in cis-(Z)- or trans-(E)-form.

Accordingly, as used herein a compound of the present invention can bein the form of one of the possible isomers, rotamers, atropisomers,tautomers or mixtures thereof, for example, as substantially puregeometric (cis or trans) isomers, diastereomers, optical isomers(antipodes), racemates or mixtures thereof.

Any resulting mixtures of isomers can be separated on the basis of thephysicochemical differences of the constituents, into the pure orsubstantially pure geometric or optical isomers, diastereomers,racemates, for example, by chromatography and/or fractionalcrystallization.

Any resulting racemates of final products or intermediates can beresolved into the optical antipodes by known methods, e.g., byseparation of the diastereomeric salts thereof, obtained with anoptically active acid or base, and liberating the optically activeacidic or basic compound. In particular, a basic moiety may thus beemployed to resolve the compounds of the present invention into theiroptical antipodes, e.g., by fractional crystallization of a salt formedwith an optically active acid, e.g., tartaric acid, dibenzoyl tartaricacid, diacetyl tartaric acid, di-O,O′-p-toluoyl tartaric acid, mandelicacid, malic acid or camphor-10-sulfonic acid. Racemic products can alsobe resolved by chiral chromatography, e.g., high pressure liquidchromatography (HPLC) using a chiral adsorbent.

As used herein, the term “pharmaceutically acceptable salts” refers tosalts that retain the biological effectiveness and properties of thecompounds of this invention and, which typically are not biologically orotherwise undesirable. In many cases, the compounds of the presentinvention are capable of forming acid and/or base salts by virtue of thepresence of amino and/or carboxyl groups or groups similar thereto.

Pharmaceutically acceptable acid addition salts can be formed withinorganic acids and organic acids, e.g., acetate, aspartate, benzoate,besylate, bromide/hydrobromide, bicarbonate/carbonate,bisulfate/sulfate, camphorsulformate, chloride/hydrochloride,chlortheophyllonate, citrate, ethandisulfonate, fumarate, gluceptate,gluconate, glucuronate, hippurate, hydroiodide/iodide, isethionate,lactate, lactobionate, laurylsulfate, malate, maleate, malonate,mandelate, mesylate, methylsulphate, naphthoate, napsylate, nicotinate,nitrate, octadecanoate, oleate, oxalate, palmitate, pamoate,phosphate/hydrogen phosphate/dihydrogen phosphate, polygalacturonate,propionate, stearate, succinate, sulfosalicylate, tartrate, tosylate andtrifluoroacetate salts.

Inorganic acids from which salts can be derived include, for example,hydrochloric acid, hydrobromic acid, sulfuric acid, nitric acid,phosphoric acid, and the like. Organic acids from which salts can bederived include, for example, acetic acid, propionic acid, glycolicacid, oxalic acid, maleic acid, malonic acid, succinic acid, fumaricacid, tartaric acid, citric acid, benzoic acid, mandelic acid,methanesulfonic acid, ethanesulfonic acid, toluenesulfonic acid,sulfosalicylic acid, and the like. Pharmaceutically acceptable baseaddition salts can be formed with inorganic and organic bases.

Inorganic bases from which salts can be derived include, for example,ammonium salts and metals from columns I to XII of the periodic table.In certain embodiments, the salts are derived from sodium, potassium,ammonium, calcium, magnesium, iron, silver, zinc, and copper;particularly suitable salts include ammonium, potassium, sodium, calciumand magnesium salts.

Organic bases from which salts can be derived include, for example,primary, secondary, and tertiary amines, substituted amines includingnaturally occurring substituted amines, cyclic amines, basic ionexchange resins, and the like. Certain organic amines includeisopropylamine, benzathine, cholinate, diethanolamine, diethylamine,lysine, meglumine, piperazine and tromethamine.

The pharmaceutically acceptable salts of the present invention can besynthesized from a parent compound, a basic or acidic moiety, byconventional chemical methods. Generally, such salts can be prepared byreacting free acid forms of these compounds with a stoichiometric amountof the appropriate base (such as Na, Ca, Mg, or K hydroxide, carbonate,bicarbonate or the like), or by reacting free base forms of thesecompounds with a stoichiometric amount of the appropriate acid. Suchreactions are typically carried out in water or in an organic solvent,or in a mixture of the two. Generally, use of non-aqueous media likeether, ethyl acetate, ethanol, isopropanol, or acetonitrile isdesirable, where practicable. Lists of additional suitable salts can befound, e.g., in “Remington's Pharmaceutical Sciences”, 20th ed., MackPublishing Company, Easton, Pa., (1985); and in “Handbook ofPharmaceutical Salts: Properties, Selection, and Use” by Stahl andWermuth (Wiley-VCH, Weinheim, Germany, 2002).

Any formula given herein is also intended to represent unlabeled formsas well as isotopically labeled forms of the compounds. For example, anyhydrogen represented by “H” in any of the formulae herein is intended torepresent all isotopic forms of hydrogen (e.g. ¹H, ²H or D, ³H); anycarbon represented by “C” in any of the formulae herein is intended torepresent all isotopic forms of carbon (e.g. ¹¹C, ¹³C, ¹⁴C); anynitrogen represented by “N” is intended to represent all isotopic formsof nitrogen (e.g. ¹⁴N, ¹⁵N). Other examples of isotopes that areincluded in the invention include isotopes of oxygen, sulfur,phosphorous, fluorine, iodine and chlorine, such as ¹⁸F ³¹P, ³²P, ³⁵S,³⁶Cl, ¹²⁵I. The invention includes various isotopically labeledcompounds as defined herein, for example those into which radioactiveisotopes, such as ³H, ¹³C, and ¹⁴C are present. In one embodiment, theatoms in the formulae herein occur in their natural abundance. Inanother embodiment, one or more hydrogen atom may be enriched in ²H;or/and one or more carbon atom may be enriched in ¹¹C, ¹³C or ¹⁴C;or/and one or more nitrogen may be enriched in ¹⁴N. Such isotopicallylabelled compounds are useful in metabolic studies (with ¹⁴C), reactionkinetic studies (with, for example ²H or ³H), detection or imagingtechniques, such as positron emission tomography (PET) or single-photonemission computed tomography (SPECT) including drug or substrate tissuedistribution assays, or in radioactive treatment of patients. Inparticular, an ¹⁸F or labeled compound may be particularly desirable forPET or SPECT studies. Isotopically labeled compounds of this inventionand prodrugs thereof can generally be prepared by carrying out theprocedures disclosed in the schemes or in the examples and preparationsdescribed below by substituting a readily available isotopically labeledreagent for a non-isotopically labeled reagent.

Further, enrichment with heavier isotopes, particularly deuterium (i.e.,²H or D) may afford certain therapeutic advantages resulting fromgreater metabolic stability, for example increased in vivo half-life orreduced dosage requirements or an improvement in therapeutic index. Itis understood that deuterium in this context is regarded as asubstituent of a compound of the formulae I to IV. The concentration ofsuch a heavier isotope, specifically deuterium, may be defined by theisotopic enrichment factor. The term “isotopic enrichment factor” asused herein means the ratio between the isotopic abundance and thenatural abundance of a specified isotope. If a substituent in a compoundof this invention is denoted deuterium, such compound has an isotopicenrichment factor for each designated deuterium atom of at least 3500(52.5% deuterium incorporation at each designated deuterium atom), atleast 4000 (60% deuterium incorporation), at least 4500 (67.5% deuteriumincorporation), at least 5000 (75% deuterium incorporation), at least5500 (82.5% deuterium incorporation), at least 6000 (90% deuteriumincorporation), at least 6333.3 (95% deuterium incorporation), at least6466.7 (97% deuterium incorporation), at least 6600 (99% deuteriumincorporation), or at least 6633.3 (99.5% deuterium incorporation).Isotopically-enriched compounds of formulae I to IV can generally beprepared by conventional techniques known to those skilled in the art orby processes analogous to those described in the accompanying Examplesand Preparations using an appropriate isotopically-enriched reagent inplace of the non-enriched reagent previously employed.

Pharmaceutically acceptable solvates in accordance with the inventioninclude those wherein the solvent of crystallization may be isotopicallysubstituted, e.g. D₂O, d₆-acetone, d₆-DMSO.

Compounds of the invention, i.e. compounds of formula I′, I, II, III orIV that contain groups capable of acting as donors and/or acceptors forhydrogen bonds may be capable of forming co-crystals with suitableco-crystal formers. These co-crystals may be prepared from compounds offormula I′, I, II, Ill, IV or V by known co-crystal forming procedures.Such procedures include grinding, heating, co-subliming, co-melting, orcontacting in solution compounds of formula I′, I, II, Ill or IV withthe co-crystal former under crystallization conditions and isolatingco-crystals thereby formed. Suitable co-crystal formers include thosedescribed in WO 2004/078163. Hence the invention further providesco-crystals comprising a compound of formula I′, I, II, III or IV, or apharmaceutically acceptable salt thereof.

As used herein, the term “pharmaceutically acceptable carrier” includesany and all solvents, dispersion media, coatings, surfactants,antioxidants, preservatives (e.g., antibacterial agents, antifungalagents), isotonic agents, absorption delaying agents, salts,preservatives, drugs, drug stabilizers, binders, excipients,disintegration agents, lubricants, sweetening agents, flavoring agents,dyes, and the like and combinations thereof, as would be known to thoseskilled in the art (see, for example, Remington's PharmaceuticalSciences, 18th Ed. Mack Printing Company, 1990, pp. 1289-1329). Exceptinsofar as any conventional carrier is incompatible with the activeingredient, its use in the therapeutic or pharmaceutical compositions iscontemplated.

The term “a therapeutically effective amount” of a compound of thepresent invention refers to an amount of the compound of the presentinvention that will elicit the biological or medical response of asubject, for example, reduction or inhibition of an enzyme or a proteinactivity, or amelioration of a symptom, alleviation of a condition, slowor delay disease progression, or prevention of a disease, etc. In onenon-limiting embodiment, the term “a therapeutically effective amount”refers to the amount of the compound of the present invention that, whenadministered to a subject, is effective to (1) at least partiallyalleviate, inhibit, prevent and/or ameliorate a condition, a disorder ora disease or a symptom thereof (i) ameliorated by the inhibition ofneutral endopeptidase EC 3.4. 24.11 or (ii) associated with neutralendopeptidase EC 3.4. 24.11 activity, or (iii) characterized by abnormalactivity of neutral endopeptidase EC 3.4. 24.11; or (2) reduce orinhibit the activity of neutral endopeptidase EC 3.4. 24.11; or (3)reduce or inhibit the expression of neutral endopeptidase EC 3.4. 24.11.In another non-limiting embodiment, the term “a therapeuticallyeffective amount” refers to the amount of the compound of the presentinvention that, when administered to a cell, or a tissue, or anon-cellular biological material, or a medium, is effective to at leastpartially reduce or inhibit the activity of neutral endopeptidase EC3.4. 24.11; or at least partially reduce or inhibit the expression ofneutral endopeptidase EC 3.4. 24.11

As used herein, the term “subject” refers to an animal. Typically theanimal is a mammal. A subject also refers to for example, primates(e.g., humans), cows, sheep, goats, horses, dogs, cats, rabbits, rats,mice, fish, birds and the like. In certain embodiments, the subject is aprimate. In yet other embodiments, the subject is a human.

As used herein, the term “inhibit”, “inhibition” or “inhibiting” refersto the reduction or suppression of a given condition, symptom, ordisorder, or disease, or a significant decrease in the baseline activityof a biological activity or process.

As used herein, the term “treat”, “treating” or “treatment” of anydisease or disorder refers in one embodiment, to ameliorating thedisease or disorder (i.e., slowing or arresting or reducing thedevelopment of the disease or at least one of the clinical symptomsthereof). In another embodiment “treat”, “treating” or “treatment”refers to alleviating or ameliorating at least one physical parameterincluding those which may not be discernible by the patient. In yetanother embodiment, “treat”, “treating” or “treatment” refers tomodulating the disease or disorder, either physically, (e.g.,stabilization of a discernible symptom), physiologically, (e.g.,stabilization of a physical parameter), or both. In yet anotherembodiment, “treat”, “treating” or “treatment” refers to preventing ordelaying the onset or development or progression of the disease ordisorder.

As used herein, a subject is “in need of” a treatment if such subjectwould benefit biologically, medically or in quality of life from suchtreatment.

As used herein, the term “a,” “an,” “the” and similar terms used in thecontext of the present invention (especially in the context of theclaims) are to be construed to cover both the singular and plural unlessotherwise indicated herein or clearly contradicted by the context.

All methods described herein can be performed in any suitable orderunless otherwise indicated herein or otherwise clearly contradicted bycontext. The use of any and all examples, or exemplary language (e.g.“such as”) provided herein is intended merely to better illuminate theinvention and does not pose a limitation on the scope of the inventionotherwise claimed.

Compounds of the present invention are either obtained in the free form,as a salt thereof, or as prodrug derivatives thereof.

When both a basic group and an acid group are present in the samemolecule, the compounds of the present invention may also form internalsalts, e.g., zwitterionic molecules.

The present invention also provides pro-drugs of the compounds of thepresent invention that converts in vivo to the compounds of the presentinvention. A pro-drug is an active or inactive compound that is modifiedchemically through in vivo physiological action, such as hydrolysis,metabolism and the like, into a compound of this invention followingadministration of the prodrug to a subject. The suitability andtechniques involved in making and using pro-drugs are well known bythose skilled in the art. Prodrugs can be conceptually divided into twonon-exclusive categories, bioprecursor prodrugs and carrier prodrugs.See The Practice of Medicinal Chemistry, Ch. 31-32 (Ed. Wermuth,Academic Press, San Diego, Calif., 2001). Generally, bioprecursorprodrugs are compounds, which are inactive or have low activity comparedto the corresponding active drug compound, that contain one or moreprotective groups and are converted to an active form by metabolism orsolvolysis. Both the active drug form and any released metabolicproducts should have acceptably low toxicity. Carrier prodrugs are drugcompounds that contain a transport moiety, e.g., that improve uptakeand/or localized delivery to a site(s) of action. Desirably for such acarrier prodrug, the linkage between the drug moiety and the transportmoiety is a covalent bond, the prodrug is inactive or less active thanthe drug compound, and any released transport moiety is acceptablynon-toxic. For prodrugs where the transport moiety is intended toenhance uptake, typically the release of the transport moiety should berapid. In other cases, it is desirable to utilize a moiety that providesslow release, e.g., certain polymers or other moieties, such ascyclodextrins. Carrier prodrugs can, for example, be used to improve oneor more of the following properties: increased lipophilicity, increasedduration of pharmacological effects, increased site-specificity,decreased toxicity and adverse reactions, and/or improvement in drugformulation (e.g., stability, water solubility, suppression of anundesirable organoleptic or physiochemical property). For example,lipophilicity can be increased by esterification of (a) hydroxyl groupswith lipophilic carboxylic acids (e.g., a carboxylic acid having atleast one lipophilic moiety), or (b) carboxylic acid groups withlipophilic alcohols (e.g., an alcohol having at least one lipophilicmoiety, for example aliphatic alcohols).

Exemplary prodrugs are, e.g., esters of free carboxylic acids and S-acylderivatives of thiols and O-acyl derivatives of alcohols or phenols,wherein acyl has a meaning as defined herein. Suitable prodrugs areoften pharmaceutically acceptable ester derivatives convertible bysolvolysis under physiological conditions to the parent carboxylic acid,e.g., lower alkyl esters, cycloalkyl esters, lower alkenyl esters,benzyl esters, mono- or di-substituted lower alkyl esters, such as theω-(amino, mono- or di-lower alkylamino, carboxy, loweralkoxycarbonyl)-lower alkyl esters, the α-(lower alkanoyloxy, loweralkoxycarbonyl or di-lower alkylaminocarbonyl)-lower alkyl esters, suchas the pivaloyloxymethyl ester and the like conventionally used in theart. In addition, amines have been masked as arylcarbonyloxymethylsubstituted derivatives which are cleaved by esterases in vivo releasingthe free drug and formaldehyde (Bundgaard, J. Med. Chem. 2503 (1989)).Moreover, drugs containing an acidic NH group, such as imidazole, imide,indole and the like, have been masked with N-acyloxymethyl groups(Bundgaard, Design of Prodrugs, Elsevier (1985)). Hydroxy groups havebeen masked as esters and ethers. EP 039,051 (Sloan and Little)discloses Mannich-base hydroxamic acid prodrugs, their preparation anduse.

Furthermore, the compounds of the present invention, including theirsalts, can also be obtained in the form of their hydrates, or includeother solvents used for their crystallization.

General Synthetic Scheme:

The compounds of the invention can be synthesized using the methodsdescribed in the following schemes, examples, and by using artrecognized techniques. All compounds described herein are included inthe invention as compounds. Compounds of the invention may besynthesized according to at least one of the methods described inschemes 1-3.

Within the scope of this text, only a readily removable group that isnot a constituent of the particular desired end product of the compoundsof the present invention is designated a “protecting group”, unless thecontext indicates otherwise. The protection of functional groups by suchprotecting groups, the protecting groups themselves, and their cleavagereactions are described for example in standard reference works, such asJ. F. W. McOmie, “Protective Groups in Organic Chemistry”, Plenum Press,London and New York 1973, in T. W. Greene and P. G. M. Wuts, “ProtectiveGroups in Organic Synthesis”, Third edition, Wiley, New York 1999.

Salts of compounds of the present invention having at least onesalt-forming group may be prepared in a manner known per se. Forexample, salts of compounds of the present invention having acid groupsmay be formed, for example, by treating the compounds with metalcompounds, such as alkali metal salts of suitable organic carboxylicacids, e.g. the sodium salt of 2-ethylhexanoic acid, with organic alkalimetal or alkaline earth metal compounds, such as the correspondinghydroxides, carbonates or hydrogen carbonates, such as sodium orpotassium hydroxide, carbonate or hydrogen carbonate, with correspondingcalcium compounds or with ammonia or a suitable organic amine,stoichiometric amounts or only a small excess of the salt-forming agentpreferably being used. Acid addition salts of compounds of the presentinvention are obtained in customary manner, e.g. by treating thecompounds with an acid or a suitable anion exchange reagent. Internalsalts of compounds of the present invention containing acid and basicsalt-forming groups, e.g. a free carboxy group and a free amino group,may be formed, e.g. by the neutralisation of salts, such as acidaddition salts, to the isoelectric point, e.g. with weak bases, or bytreatment with ion exchangers.

Salts can be converted in customary manner into the free compounds;metal and ammonium salts can be converted, for example, by treatmentwith suitable acids, and acid addition salts, for example, by treatmentwith a suitable basic agent.

Mixtures of isomers obtainable according to the invention can beseparated in a manner known per se into the individual isomers;diastereoisomers can be separated, for example, by partitioning betweenpolyphasic solvent mixtures, recrystallisation and/or chromatographicseparation, for example over silica gel or by e.g. medium pressureliquid chromatography over a reversed phase column, and racemates can beseparated, for example, by the formation of salts with optically puresalt-forming reagents and separation of the mixture of diastereoisomersso obtainable, for example by means of fractional crystallisation, or bychromatography over optically active column materials.

Intermediates and final products can be worked up and/or purifiedaccording to standard methods, e.g. using chromatographic methods,distribution methods, (re-) crystallization, and the like.

The following applies in general to all processes mentioned hereinbefore and hereinafter.

All the above-mentioned process steps can be carried out under reactionconditions that are known per se, including those mentionedspecifically, in the absence or, customarily, in the presence ofsolvents or diluents, including, for example, solvents or diluents thatare inert towards the reagents used and dissolve them, in the absence orpresence of catalysts, condensation or neutralizing agents, for exampleion exchangers, such as cation exchangers, e.g. in the H+ form,depending on the nature of the reaction and/or of the reactants atreduced, normal or elevated temperature, for example in a temperaturerange of from about −100° C. to about 190° C., including, for example,from approximately −80° C. to approximately 150° C., for example at from−80 to −60° C., at room temperature, at from −20 to 40° C. or at refluxtemperature, under atmospheric pressure or in a closed vessel, whereappropriate under pressure, and/or in an inert atmosphere, for exampleunder an argon or nitrogen atmosphere.

At all stages of the reactions, mixtures of isomers that are formed canbe separated into the individual isomers, for example diastereoisomersor enantiomers, or into any desired mixtures of isomers, for exampleracemates or mixtures of diastereoisomers, for example analogously tothe methods described under “Additional process steps”.

The solvents from which those solvents that are suitable for anyparticular reaction may be selected include those mentioned specificallyor, for example, water, esters, such as lower alkyl-lower alkanoates,for example ethyl acetate, ethers, such as aliphatic ethers, for examplediethyl ether, or cyclic ethers, for example tetrahydrofuran or dioxane,liquid aromatic hydrocarbons, such as benzene or toluene, alcohols, suchas methanol, ethanol or 1- or 2-propanol, nitriles, such asacetonitrile, halogenated hydrocarbons, such as methylene chloride orchloroform, acid amides, such as dimethylformamide or dimethylacetamide, bases, such as heterocyclic nitrogen bases, for examplepyridine or N-methylpyrrolidin-2-one, carboxylic acid anhydrides, suchas lower alkanoic acid anhydrides, for example acetic anhydride, cyclic,linear or branched hydrocarbons, such as cyclohexane, hexane orisopentane, methycyclohexane, or mixtures of those solvents, for exampleaqueous solutions, unless otherwise indicated in the description of theprocesses. Such solvent mixtures may also be used in working up, forexample by chromatography or partitioning.

The compounds, including their salts, may also be obtained in the formof hydrates, or their crystals may, for example, include the solventused for crystallization. Different crystalline forms may be present.

The invention relates also to those forms of the process in which acompound obtainable as an intermediate at any stage of the process isused as starting material and the remaining process steps are carriedout, or in which a starting material is formed under the reactionconditions or is used in the form of a derivative, for example in aprotected form or in the form of a salt, or a compound obtainable by theprocess according to the invention is produced under the processconditions and processed further in situ.

All starting materials, building blocks, reagents, acids, bases,dehydrating agents, solvents and catalysts utilized to synthesize thecompounds of the present invention are either commercially available orcan be produced by organic synthesis methods known to one of ordinaryskill in the art (Houben-Weyl 4^(th) Ed. 1952, Methods of OrganicSynthesis, Thieme, Volume 21).

Typically, the compounds according to of formulae I to IV can beprepared according to the Schemes 1 to 7 provided infra.

The compounds of the invention of Formula I′ or I wherein X¹ is hydroxycan be prepared by hydrolysis of intermediate A wherein A¹, R¹, R², R³,R⁴ and m have the definition of Formula I′ or I, supra; and P¹ is anappropriate protecting groups selected from, but not limited to, methyl,ethyl, or tert-butyl, or methoxybenzyl, or benzyl.

Standard methods can be applied for the hydrolysis of the intermediate Ausing a base selected from, but not limited to, NaOH, KOH, or LiOH, oran acid selected from, but not limited to, TFA, HCl, or BCl₃. When P¹ isbenzyl or methoxybenzyl, a preferable method of the deprotection ishydrogenation in the presence of a catalyst such as, but not limited to,palladium-on-carbon.

In some cases, the hydrolysis of intermediate A is not required; forexample when intermediate A is a compound of the invention of Formula I′or I wherein X¹ is O-alkyl.

Scheme 1 illustrates the synthesis of intermediate A. The intermediate Acan be prepared according to the following general procedures describedin Scheme 1 wherein A¹, P¹, R¹, R², R³, R⁴, R⁵, R⁶, s and m are aspreviously defined.

In step 1a, the intermediate 3A can be prepared by cross-coupling of anintermediate 1A wherein P² is an appropriate protecting groups selectedfrom, but not limited to, t-butoxycarbony, benzyloxycarbony,fluorenylmethyloxycarbonyl, benzyl, or methoxybenzyl and wherein LG¹ isa leaving group selected from, but not limited to, halo (e.g. bromo,chloro, or iodo) or trifluoromethanesulfonyloxy with an intermediate 2Awherein R² and m are as previously described and wherein BG is anappropriate groups selected from, but not limited to, boronic acid,trifluoroborate or boronic ester. Known coupling methods may be appliedincluding Suzuki-Miyaura coupling of the intermediate 1A with theintermediate 2A using palladium species such as, but not limited to,Pd(PPh₃)₄, PdCl₂(dppf), Pd(PPh₃)₂Cl₂, or Pd(OAc)₂ with a phosphineligand such as PPh₃, dppf, PCy₃, or P(t-Bu)₃ and a base such as, but notlimited to, Na₂CO₃, K₃PO₄, K₂CO₃, KF, CsF, NaO-t-Bu, or KO-t-Bu.

In step 2a, the intermediate 4A can be prepared by appropriateprotection of an intermediate 3A wherein P³ is a protection group suchas, but not limited to, t-butyl, methyl, benzyl, fluorenylmethyl, allylor methoxybenzyl; followed by an appropriate deprotection of the P²group.

In step 3a, the intermediate 5A can be prepared by reacting anintermediate 4A wherein R², R⁵, R⁶, s, m, and P³ are as previouslydefined with an intermediate 6A wherein R¹ and P¹ are as previouslydefined above and wherein LG² is a leaving group selected from, but notlimited to, trifluoromethansulfonyloxy, toluenesulfonyloxy,methansulfonyloxy, iodo, bromo, and chloro, followed by deprotection ofthe P³ using an appropriate method. Alternatively, the intermediates 5Acan be prepared by reacting an intermediate 4A with an intermediate 6Bwherein R¹ and P¹ are as defined above, followed by deprotection of theP³ using an appropriate method. Known coupling methods may be appliedincluding alkylation of the intermediate 4A with the intermediate 6Ausing a base such as, but not limited to, tertiary amine (e.g.triethylamine or N,N-diisopropyl ethylamine), pyridine, or K₂CO₃; orreductive amination condition of intermediate 4A with the intermediate6B, under condition such as hydrogenation in the presence of a catalystsuch as palladium-on-carbon or reduction using a reductive reagent (e.g.NaBH₄, NaBH(OAc)₃, or NaBH₃CN) in the presence of or absence of an acidsuch as acetic acid, TFA, or Ti(i-PrO)₄.

In step 4a, the intermediate A can be prepared by coupling anintermediate 5A wherein P¹, R¹, R², R⁵, R⁶, s and m are as previouslydescribed with an intermediate 7A wherein A¹, R³, and R⁴ are previouslydescribed. Known coupling methods may be applied including, but notlimited to, conversion of the intermediate 5A to a correspondingoxazolidine-2,5-dione, using reagents such as triphosgene,carbonyldiimidazole, 4-nitrophenyl chloroformate, or disuccinimidylcarbonate, conversion of the intermediate 5A to a corresponding acidhalide, using reagents such as thionyl chloride or oxalyl chloride, orconversion of the intermediate 5A to a corresponding mixed anhydrideusing reagents such as ClC(O)O-isobutyl, 2,4,6-trichlorobenzoyl chlorideor propyl phosphonic acid anhydride cyclic trimer (T3P), followed byreaction of the oxazolidine-2,5-dione, the acid halide, or the mixedanhydride with the intermediate 7A in a presence or absence of a basesuch as tertiary amine (e.g. triethylamine or N,N-diisoproplylethylamine) or K₂CO₃. Alternatively, the intermediate 5A can be coupledwith the intermediate 7A using peptide condensation reagents including,but not limited to, dicyclohexylcarbodiimide (DCC),diisopropylcarbodiimide (DIC),1-ethyl-3-(3-dimethyllaminopropyl)carbodiimide hydrochloride (EDC HCl),benzotriazole-1-yl-oxy-tris-pyrrolidino-phosphonium hexafluorophosphate(PyBOP), or benzotriazole-1-yl-oxy-tris-(dimethylamino)-phosphoniumhexafluorophosphate (BOP) in presence of or absence of a reagent such as1-hydroxybenzotriazole, 1-hydroxy-7-azabenzotriazole, ordimethylaminopyridine.

Scheme 2 illustrates the synthesis of Intermediate 5A. The intermediate5A can also be prepared according to the following procedures describedin Scheme 2 wherein BG, LG¹, LG², P¹, P³, R¹, R², and m are aspreviously defined.

in step 1b, the intermediate 9A can be prepared by reacting anintermediate 8A where in LG¹, R⁵, R⁶, s and P³ are previously describedwith an intermediate 6A wherein R¹, P¹, and LG² are as previouslydescribed, followed by an appropriate deprotection of the protectinggroup P³. Alternatively, the intermediates 9A can be prepared byreacting an intermediate 8A with an intermediate 6B wherein P¹ and R¹are as previously described, followed by an appropriate deprotection ofthe protecting group P³. Known reaction methods may be applied includingalkylation of the intermediate 8A with the intermediate 6A using a basesuch as, but not limited to, tertiary amine (e.g. triethylamine orN,N-diisoproplyl ethylamine), pyridine, or K₂CO₃, or reductive aminationcondition of intermediate 8A with the intermediate 6B, under conditionsuch as hydrogenation in the presence of a catalyst such aspalladium-on-carbon or reduction using a reducing reagent (e.g. NaBH₄,NaBH(OAc)₃, or NaBH₃CN) in the presence of or absence of an acid such asacetic acid, TFA, or Ti(i-PrO)₄.

In step 2b, the intermediate 5A can be prepared by cross-coupling of anintermediate 9A wherein LG¹, P¹, R⁵, R⁶, R¹ and s with an intermediate2A wherein BG, m, and R² are as previously described. Known couplingmethods may be applied including Suzuki-Miyaura coupling of theintermediate 9A with the intermediate 2A using palladium species suchas, but not limited to, Pd(PPh₃)₄, PdCl₂(dppf), or Pd(OAc)₂ with aphosphine ligand such as PPh₃, dppf, PCy₃, or P(t-Bu)₃ and a base suchas, but not limited to, Na₂CO₃, K₃PO₄, K₂CO₃, KF, CsF, NaO-t-Bu, orKO-t-Bu.

The intermediates 9A can also be prepared according to the followinggeneral procedure described in Scheme 3 wherein LG¹, P¹, P³, R⁵, R⁶, R¹and s are as previously described.

In step 1c, the intermediate 9A can be prepared by reductive aminationof the intermediate 10A wherein LG¹, R⁶, s and P³ are as previouslydescribed with the intermediate 11A wherein P¹, R⁵ and R¹ are aspreviously described. Known reductive amination methods may be appliedincluding a condition such as, but not limited to, hydrogenation in thepresence of a catalyst such as palladium-on-carbon or reduction using areagent such as, but not limited to, NaBH₄, NaBH(OAc)₃, or NaBH₃CN inthe presence of or absence of an acid such as acetic acid, TFA, orTi(i-PrO)₄. The intermediate 10A can be prepared according to thereported procedure. The illustrative example of this chemistry isoutlined in WO 2006015885.

The intermediate 5A can also be prepared according to the followinggeneral procedures described in Scheme 4 wherein m, P¹, P³, R¹, and R²are as previously described.

In step 1d, the intermediate 5A can be prepared by reductive aminationof the intermediate 12A wherein m, P³, R⁵, R⁶, s, m and R² are aspreviously described with the intermediate 11A wherein P′, R⁵ and R¹ areas previously described. Known reductive amination methods may beapplied including a condition such as, but not limited to, hydrogenationin the presence of a catalyst such as palladium-on-carbon or reductionusing a reagent such as, but not limited to, NaBH₄, NaBH(OAc)₃, orNaBH₃CN in the presence of or absence of an acid such as acetic acid,TFA, or Ti(i-PrO)₄. The intermediates 12A can be prepared according tothe reported procedure. The illustrative example of this chemistry isoutlined in WO 2006015885.

The intermediate A can also be prepared according to the followingprocedures described in Scheme 5 wherein A¹, LG², P¹, P², R¹, R², R³,R⁴, R⁵, R⁶, s and m are as previously described.

In step 1e, the intermediate 13A can be prepared by coupling anintermediate 3A with an intermediate 7A. Known coupling methods may beapplied including, but not limited to, conversion of the intermediate 3Ato a corresponding oxazolidine-2,5-dione, using reagents such astriphosgene, carbonyldiimidazole, 4-nitrophenyl chloroformate, ordisuccinimidyl carbonate, conversion of the intermediate 3A to acorresponding acid halide, using reagents such as thionyl chloride oroxalyl chloride, or conversion of the intermediate 3A to a correspondingmixed anhydride using reagents such as ClC(O)O-isobutyl or2,4,6-trichlorobenzoyl chloride, followed by reaction of theoxazolidine-2,5-dione, the acid halide, or the mixed anhydride with theintermediate 7A in a presence or absence of a base such as tertiaryamine (e.g. triethylamine or N,N-diisoproplyl ethylamine) or K₂CO₃ andan appropriate deprotection of P² protecting group. Alternatively, theintermediate 3A can be coupled with the intermediate 7A using peptidecondensation reagents including, but not limited to,dicyclohexylcarbodiimide (DCC), diisopropylcarbodiimide (DIC),1-ethyl-3-(3-dimethyllaminopropyl)carbodiimide hydrochloride (EDC HCl),benzotriazole-1-yl-oxy-tris-pyrrolidino-phosphonium hexafluorophosphate(PyBOP), or benzotriazole-1-yl-oxy-tris-(dimethylamino)-phosphoniumhexafluorophosphate (BOP) in presence of or absence of a reagent such as1-hydroxybenzotriazole, 1-hydroxy-7-azabenzotriazole, ordimethylaminopyridine followed by an appropriate deprotection of P²protecting group.

In step 2e, the intermediate A can be prepared by reacting anintermediate 13A with an intermediate 6A wherein LG² is as previouslydescribed. Alternatively, the intermediates A can be prepared byreacting an intermediate 13A with an intermediate 6B. Known reactionmethods may be applied including alkylation of the intermediate 13A withthe intermediate 6A using a base such as, but not limited to, tertiaryamine (e.g. triethylamine or NA-diisoproplylethylamine), pyridine, orK₂CO₃ or reductive amination of the intermediate 13A with theintermediate 6B under a condition such as, but not limited to,hydrogenation in the presence of a catalyst such as palladium-on-carbonor reduction using a reagent such as, but not limited to, NaBH₄,NaBH(OAc)₃, or NaBH₃CN in the presence of or absence of an acid such asacetic acid, TFA, or Ti(i-PrO)₄.

The intermediates A can also be prepared according to the followingprocedures described in Scheme 6 wherein A¹, BG, LG¹, P¹, R¹, R², R³,R⁴, R⁵, R⁶, s and m are as previously described.

In step 1f, an intermediate 14A can be prepared by coupling theintermediate 9A wherein LG¹, P¹, R⁵, R⁶, m, s and R¹ are as previouslydescribed with an intermediate 7A. Known coupling methods may be appliedincluding, but not limited to, conversion of the intermediate 9A to acorresponding oxazolidine-2,5-dione, using reagents such as triphosgene,carbonyldiimidazole, 4-nitrophenyl chloroformate, or disuccinimidylcarbonate, conversion of the intermediate 9A to a corresponding acidhalide, using reagents such as thionyl chloride or oxalyl chloride, orconversion of the intermediate 9A to a corresponding mixed anhydrideusing reagents such as ClC(O)O-isobutyl or 2,4,6-trichlorobenzoylchloride, followed by reaction of the oxazolidine-2,5-dione, the acidhalide, or the mixed anhydride with the intermediate 7A in a presence orabsence of a base such as tertiary amine (e.g. triethylamine orN,N-diisoproplylethylamine) or K₂CO₃. Alternatively, the intermediate 9Acan be coupled with the intermediate 7A using peptide condensationreagents including, but not limited to, dicyclohexylcarbodiimide (DCC),diisopropylcarbodiimide (DC),1-ethyl-3-(3-dimethyllaminopropyl)carbodiimide hydrochloride (EDC HCl),benzotriazole-1-yl-oxy-tris-pyrrolidino-phosphonium hexafluorophosphate(PyBOP), or benzotriazole-1-yl-oxy-tris-(dimethylamino)-phosphoniumhexafluorophosphate (BOP) in presence of or absence of a reagent such as1-hydroxybenzotriazole, 1-hydroxy-7-azabenzotriazole, ordimethylaminopyridine.

In step 2f, the intermediate A can be prepared by cross-coupling of anintermediate 14A wherein A¹, LG¹, P¹, R¹, R³, R⁵, R⁶, m, s and R⁴ are aspreviously described with an intermediate 2A wherein R², m, and BG areas previously described. Known coupling methods may be applied includingSuzuki-Miyaura coupling of the intermediate 14A with the intermediate 2Ausing palladium species such as, but not limited to, Pd(PPh₃)₄,PdCl₂(dppf), or Pd(OAc)₂ with a phosphine ligand such as PPh₃, dppf,PCy₃, or P(t-Bu)₃ and a base such as, but not limited to, Na₂CO₃, K₃PO₄,K₂CO₃, KF, CsF, NaO-t-Bu, or KO-t-Bu.

The intermediates 14A can also be prepared according to the followingprocedures described in Scheme 7 wherein A¹, LG¹, LG², P¹, P², R¹, R²,R³, R⁴, R⁵, R⁶, s and m are as previously described.

In step 1g, an intermediate 15A can be prepared by coupling theintermediate 1A wherein P², R⁵, R⁶, s and LG¹ are as previouslydescribed with an intermediate 7A wherein A¹, R³, and R⁴ are aspreviously described followed by an appropriate deprotection of theprotecting group P². Known coupling methods may be applied including,but not limited to, conversion of the intermediate 1A to correspondingoxazolidine-2,5-dione, using reagents such as triphosgene,carbonyldiimidazole, 4-nitrophenyl chloroformate, or disuccinimidylcarbonate, conversion of the intermediate 1A to corresponding acidhalide, using reagents such as thionyl chloride or oxalyl chloride, orconversion of the intermediate 1A to corresponding mixed anhydride usingreagents such as ClC(O)O-isobutyl or 2,4,6-trichlorobenzoyl chloride,followed by reaction of the oxazolidine-2,5-dione, the acid halide, orthe mixed anhydride with the intermediate 7A in a presence or absence ofa base such as tertiary amine (e.g. triethylamine or N,N-diisoproplylethylamine) or K₂CO₃. Alternatively, the intermediate 1A can be coupledwith the intermediate 7A using peptide condensation reagents including,but not limited to, dicyclohexylcarbodiimide (DCC),diisopropylcarbodiimide (DIC),1-ethyl-3-(3-dimethyllaminopropyl)carbodiimide hydrochloride (EDC HCl),benzotriazole-1-yl-oxy-tris-pyrrolidino-phosphonium hexafluorophosphate(PyBOP), or benzotriazole-1-yl-oxy-tris-(dimethylamino)-phosphoniumhexafluorophosphate (BOP) in presence of or absence of a reagent such as1-hydroxybenzotriazole, 1-hydroxy-7-azabenzotriazole, ordimethylaminopyridine.

In step 2g, the intermediate 14A can be prepared by reacting anintermediate 15A wherein A¹, LW, R³, R⁵, R⁶, s and R⁴ are as previouslydefined with an intermediate 6A wherein R¹, P¹, and LG² are aspreviously defined. Alternatively, the intermediates 14A can be preparedby reacting an intermediate 15A wherein A¹, LG¹, R³, R⁵, R⁶, s and R⁴are as previously defined with an intermediate 6B wherein R¹ and P¹ areas previously described. Known reaction methods may be applied includingalkylation of the intermediate 15A with the intermediate 6A using a basesuch as, but not limited to, tertiary amine (e.g. triethylamine orN,N-diisoproplyl ethylamine), pyridine, or K₂CO₃ or reductive aminationof the intermediate 15A with the intermediate 6B under a condition suchas, but not limited to, hydrogenation in the presence of a catalyst suchas palladium-on-carbon or reduction using a reagent such as, but notlimited to, NaBH₄, NaBH(OAc)₃, or NaBH₃CN in the presence of or absenceof an acid such as acetic acid, TFA, or Ti(i-PrO)₄.

The invention further includes any variant of the present processes, inwhich an intermediate product obtainable at any stage thereof is used asstarting material and the remaining steps are carried out, or in whichthe starting materials are formed in situ under the reaction conditions,or in which the reaction components are used in the form of their saltsor optically pure antipodes.

Compounds of the invention and intermediates can also be converted intoeach other according to methods generally known to those skilled in theart.

In another aspect, the present invention provides a pharmaceuticalcomposition comprising a compound of the present invention or apharmaceutically acceptable salt thereof and one or morepharmaceutically acceptable carriers. The pharmaceutical composition canbe formulated for particular routes of administration such as oraladministration, parenteral administration, and rectal administration,etc. In addition, the pharmaceutical compositions of the presentinvention can be made up in a solid form (including without limitationcapsules, tablets, pills, granules, powders or suppositories), or in aliquid form (including without limitation solutions, suspensions oremulsions). The pharmaceutical compositions can be subjected toconventional pharmaceutical operations such as sterilization and/or cancontain conventional inert diluents, lubricating agents, or bufferingagents, as well as adjuvants, such as preservatives, stabilizers,wetting agents, emulsifers and buffers, etc.

Typically, the pharmaceutical compositions are tablets or gelatincapsules comprising the active ingredient together with

-   -   a) diluents, e.g., lactose, dextrose, sucrose, mannitol,        sorbitol, cellulose and/or glycine;    -   b) lubricants, e.g., silica, talcum, stearic acid, its magnesium        or calcium salt and/or polyethyleneglycol; for tablets also    -   c) binders, e.g., magnesium aluminum silicate, starch paste,        gelatin, tragacanth, methylcellulose, sodium        carboxymethylcellulose and/or polyvinylpyrrolidone; if desired    -   d) disintegrants, e.g., starches, agar, alginic acid or its        sodium salt, or effervescent mixtures; and/or    -   e) absorbents, colorants, flavors and sweeteners.

Tablets may be either film coated or enteric coated according to methodsknown in the art.

Suitable compositions for oral administration include an effectiveamount of a compound of the invention in the form of tablets, lozenges,aqueous or oily suspensions, dispersible powders or granules, emulsion,hard or soft capsules, or syrups or elixirs. Compositions intended fororal use are prepared according to any method known in the art for themanufacture of pharmaceutical compositions and such compositions cancontain one or more agents selected from the group consisting ofsweetening agents, flavoring agents, coloring agents and preservingagents in order to provide pharmaceutically elegant and palatablepreparations. Tablets may contain the active ingredient in admixturewith nontoxic pharmaceutically acceptable excipients which are suitablefor the manufacture of tablets. These excipients are, for example, inertdiluents, such as calcium carbonate, sodium carbonate, lactose, calciumphosphate or sodium phosphate; granulating and disintegrating agents,for example, corn starch, or alginic acid; binding agents, for example,starch, gelatin or acacia; and lubricating agents, for example magnesiumstearate, stearic acid or talc. The tablets are uncoated or coated byknown techniques to delay disintegration and absorption in thegastrointestinal tract and thereby provide a sustained action over alonger period. For example, a time delay material such as glycerylmonostearate or glyceryl distearate can be employed. Formulations fororal use can be presented as hard gelatin capsules wherein the activeingredient is mixed with an inert solid diluent, for example, calciumcarbonate, calcium phosphate or kaolin, or as soft gelatin capsuleswherein the active ingredient is mixed with water or an oil medium, forexample, peanut oil, liquid paraffin or olive oil.

Certain injectable compositions are aqueous isotonic solutions orsuspensions, and suppositories are advantageously prepared from fattyemulsions or suspensions. Said compositions may be sterilized and/orcontain adjuvants, such as preserving, stabilizing, wetting oremulsifying agents, solution promoters, salts for regulating the osmoticpressure and/or buffers. In addition, they may also contain othertherapeutically valuable substances. Said compositions are preparedaccording to conventional mixing, granulating or coating methods,respectively, and contain about 0.1-75%, or contain about 1-50%, of theactive ingredient.

Suitable compositions for transdermal application include an effectiveamount of a compound of the invention with a suitable carrier. Carrierssuitable for transdermal delivery include absorbable pharmacologicallyacceptable solvents to assist passage through the skin of the host. Forexample, transdermal devices are in the form of a bandage comprising abacking member, a reservoir containing the compound optionally withcarriers, optionally a rate controlling barrier to deliver the compoundof the skin of the host at a controlled and predetermined rate over aprolonged period of time, and means to secure the device to the skin.

Suitable compositions for topical application, e.g., to the skin andeyes, include aqueous solutions, suspensions, ointments, creams, gels orsprayable formulations, e.g., for delivery by aerosol or the like. Suchtopical delivery systems will in particular be appropriate for dermalapplication. They are thus particularly suited for use in topical,including cosmetic, formulations well-known in the art. Such may containsolubilizers, stabilizers, tonicity enhancing agents, buffers andpreservatives.

As used herein a topical application may also pertain to an inhalationor to an intranasal application. They may be conveniently delivered inthe form of a dry powder (either alone, as a mixture, for example a dryblend with lactose, or a mixed component particle, for example withphospholipids) from a dry powder inhaler or an aerosol spraypresentation from a pressurised container, pump, spray, atomizer ornebuliser, with or without the use of a suitable propellant.

The present invention further provides anhydrous pharmaceuticalcompositions and dosage forms comprising the compounds of the presentinvention as active ingredients, since water may facilitate thedegradation of certain compounds.

Anhydrous pharmaceutical compositions and dosage forms of the inventioncan be prepared using anhydrous or low moisture containing ingredientsand low moisture or low humidity conditions. An anhydrous pharmaceuticalcomposition may be prepared and stored such that its anhydrous nature ismaintained. Accordingly, anhydrous compositions are packaged usingmaterials known to prevent exposure to water such that they can beincluded in suitable formulary kits. Examples of suitable packaginginclude, but are not limited to, hermetically sealed foils, plastics,unit dose containers (e.g., vials), blister packs, and strip packs.

The invention further provides pharmaceutical compositions and dosageforms that comprise one or more agents that reduce the rate by which thecompound of the present invention as an active ingredient willdecompose. Such agents, which are referred to herein as “stabilizers,”include, but are not limited to, antioxidants such as ascorbic acid, pHbuffers, or salt buffers, etc.

The compounds according to anyone of formulae I to IV, or apharmaceutically acceptable salt thereof, in free form or inpharmaceutically acceptable salt form, exhibit valuable pharmacologicalproperties, e.g. neutral endopeptidase EC 3.4. 24.11 modulatingproperties, e.g. as indicated in in vitro and in vivo tests as providedin the next sections and are therefore indicated for therapy.

Compounds of the invention or a pharmaceutically acceptable saltthereof, may be useful in the treatment of an indication selected fromhypertension, resistant hypertension, pulmonary hypertension, pulmonaryarterial hypertension, isolated systolic hypertension, peripheralvascular disease, heart failure, congestive heart failure, leftventricular hypertrophy, angina, renal insufficiency (diabetic ornon-diabetic), renal failure (including edema and salt retension),diabetic nephropathy, non-diabetic nephropathy, nephroic syndrome,glomerulonephritis, scleroderma, glomerular sclerosis, proteinurea ofprimary renal disease, renal vascular hypertention, diabetic retinopathyand end-stage renal disease (ESRD), endothelial dysfunction, diastolicdysfunction, hypertrophic cardiomyopathy, diabetic cardiac myopathy,supraventricular and ventricular arrhythmias, atrial fibrillation (AF),cardiac fibrosis, atrial flutter, detrimental vascular remodeling,plaque stabilization, myocardial infarction (MI), renal fibrosis,polycystic kidney disease (PKD), renal failure (including edema and saltretension), cyclical oedema, Menières disease, hyperaldosteroneism(primary and secondary) and hypercalciuria, ascites, glaucoma, menstrualdisorders, preterm labour, pre-eclampsia, endometriosis, andreproductive disorders (especially male and female infertility,polycystic ovarian syndrome, implantation failure), asthma, obstructivesleep apnea, inflammation, leukemia, pain, epilepsy, affective disorderssuch as depression and psychotic condition such as dementia andgeriatric confusion, obesity and gastrointestinal disorders (especiallydiarrhea and irritable bowel syndrome), wound healing (especiallydiabetic and venous ulcers and pressure sores), septic shock, gastricacid secretion dysfunctions, hyperreninaemia, cystic fibrosis,restenosis, type-2 diabetes, metabolic syndrome, diabetic complicationsand atherosclerosis, male and female sexual dysfunction. Thus, as afurther embodiment, the present invention provides the use of a compoundof formula I′, I, II, Ill or IV, or a pharmaceutically acceptable saltthereof. In a further embodiment, the therapy is selected from a diseasewhich is associated with neutral endopeptidase EC 3.4. 24.11 activity.In another embodiment, the disease is selected from the afore-mentionedlist, suitably hypertension, resistant hypertension, pulmonaryhypertension, pulmonary arterial hypertension, isolated systolichypertension, peripheral vascular disease, heart failure, congestiveheart failure, left ventricular hypertrophy, angina, renalinsufficiency, renal failure (including edema and salt retension),diabetic nephropathy, non-diabetic nephropathy, type-2 diabetes, anddiabetic complications and most suitably cardiovascular disorders, suchas hypertension, renal insufficiency including edema and congestiveheart failure.

Thus, as a further embodiment, the present invention provides the use ofa compound of formula I′ I, II, Ill or IV, or a pharmaceuticallyacceptable salt thereof, in therapy. In a further embodiment, thetherapy is selected from a disease which may be treated by inhibitingneutral endopeptidase EC. 3.4. 24.11. activity.

In another embodiment, the invention provides a method of treating adisease which is associated with neutral endopeptidase EC 3.4. 24.11activity comprising administration of a therapeutically acceptableamount of a compound of formula I′, I, II, Ill or IV, or apharmaceutically acceptable salt thereof. In a further embodiment, thedisease is selected from the afore-mentioned list, suitablyhypertension, resistant hypertension, pulmonary hypertension, pulmonaryarterial hypertension, isolated systolic hypertension, peripheralvascular disease, heart failure, congestive heart failure, leftventricular hypertrophy, angina, renal insufficiency, renal failure(including edema and salt retension), diabetic nephropathy, non-diabeticnephropathy, type-2 diabetes, and diabetic complications and mostsuitably cardiovascular disorders, such as hypertension, renalinsufficiency including edema and congestive heart failure.

The pharmaceutical composition or combination of the present inventioncan be in unit dosage of about 1-1000 mg of active ingredient(s) for asubject of about 50-70 kg, or about 1-500 mg or about 1-250 mg or about1-150 mg or about 0.5-100 mg, or about 1-50 mg of active ingredients.The therapeutically effective dosage of a compound, the pharmaceuticalcomposition, or the combinations thereof, is dependent on the species ofthe subject, the body weight, age and individual condition, the disorderor disease or the severity thereof being treated. A physician, clinicianor veterinarian of ordinary skill can readily determine the effectiveamount of each of the active ingredients necessary to prevent, treat orinhibit the progress of the disorder or disease.

The pharmaceutical composition or combination of the present inventioncan be in unit dosage of about 1-1000 mg of active ingredient(s) for asubject of about 50-70 kg, or about 1-500 mg or about 1-250 mg or about1-150 mg or about 0.5-100 mg, or about 1-50 mg of active ingredients.The therapeutically effective dosage of a compound, the pharmaceuticalcomposition, or the combinations thereof, is dependent on the species ofthe subject, the body weight, age and individual condition, the disorderor disease or the severity thereof being treated. A physician, clinicianor veterinarian of ordinary skill can readily determine the effectiveamount of each of the active ingredients necessary to prevent, treat orinhibit the progress of the disorder or disease.

The above-cited dosage properties are demonstrable in vitro and in vivotests using advantageously mammals, e.g., mice, rats, dogs, monkeys orisolated organs, tissues and preparations thereof. The compounds of thepresent invention can be applied in vitro in the form of solutions,e.g., aqueous solutions, and in vivo either enterally, parenterally,advantageously intravenously, e.g., as a suspension or in aqueoussolution. The dosage in vitro may range between about 10⁻³ molar and10⁻⁹ molar concentrations. A therapeutically effective amount in vivomay range depending on the route of administration, between about0.1-500 mg/kg, or between about 1-100 mg/kg.

The activity of a compound according to the present invention can beassessed by the following in vitro & in vivo methods and/or by thefollowing in vitro & in vivo methods well-described in the art. See Afluorescence lifetime-based assay for protease inhibitor profiling onhuman kallikrein 7 Doering K, Meder G, Hinnenberger M, Woelcke J, Mayr LM, Hassiepen U Biomol Screen. 2009 January; 14(1):1-9.

In particular, the in vitro inhibition of recombinant human neutralendopeptidase (NEP, EC 3.424.11) can be determined as follows:

Recombinant human neutral endopeptidase (expressed in insect cells andpurified using standard methods, final concentration 7 μM) ispre-incubated with test compounds at various concentrations for 1 hourat room temperature in 10 mM sodium phosphate buffer at pH 7.4,containing 150 mM NaCl and 0.05% (w/v) CHAPS. The enzymatic reaction isstarted by the addition of a synthetic peptide substrateCys(PT14)-Arg-Arg-Leu-Trp-OH to a final concentration of 0.7 μM.Substrate hydrolysis leads to an increase fluorescence lifetime (FLT) ofPT14 measured by the means of a FLT reader as described by Doering etal. (2009). The effect of the compound on the enzymatic activity wasdetermined after 1 hour (t=60 min) incubation at room temperature. TheIC50 values, corresponding to the inhibitor concentration showing 50%reduction of the FLT values measured in absence of inhibitor, arecalculated from the plot of percentage of inhibition vs. inhibitorconcentration using non-linear regression analysis software.

Using the test assay (as described above) compounds of the inventionexhibited inhibitory efficacy in accordance to Table 1, provided infra.

TABLE 1 Inhibitory Activity of Compounds Compounds: Example # Human NEPIC₅₀ (nM) Example 3-1 0.09 Example 3-2 0.3 Example 3-4 11 Example 3-72.4 Example 3-10 91 Example 3-12 0.2 Example 3-13 0.2

The compound of the present invention may be administered eithersimultaneously with, or before or after, one or more other therapeuticagent. The compound of the present invention may be administeredseparately, by the same or different route of administration, ortogether in the same pharmaceutical composition as the other agents.

In one embodiment, the invention provides a product comprising acompound of formula I′, I, II, Ill or IV, or a pharmaceuticallyacceptable salt thereof, and at least one other therapeutic agent as acombined preparation for simultaneous, separate or sequential use intherapy. In one embodiment, the therapy is the treatment of a disease orcondition associated with neutral endopeptidase EC 3.4. 24.11 activity.

Products provided as a combined preparation include a compositioncomprising the compound of formula I′, I, II, Ill or IV, or apharmaceutically acceptable salt thereof, and the other therapeuticagent(s) together in the same pharmaceutical composition, or thecompound of formula I′, I, II, III or IV, or a pharmaceuticallyacceptable salt thereof, and the other therapeutic agent(s) in separateform, e.g. in the form of a kit.

In one embodiment, the invention provides a pharmaceutical compositioncomprising a compound of formula I′, I, II, III or IV, or apharmaceutically acceptable salt thereof, and another therapeuticagent(s). Optionally, the pharmaceutical composition may comprise apharmaceutically acceptable excipient, as described above.

In one embodiment, the invention provides a kit comprising two or moreseparate pharmaceutical compositions, at least one of which contains acompound of formula I′, I, II, III or IV, or a pharmaceuticallyacceptable salt thereof. In one embodiment, the kit comprises means forseparately retaining said compositions, such as a container, dividedbottle, or divided foil packet. An example of such a kit is a blisterpack, as typically used for the packaging of tablets, capsules and thelike.

The kit of the invention may be used for administering different dosageforms, for example, oral and parenteral, for administering the separatecompositions at different dosage intervals, or for titrating theseparate compositions against one another. To assist compliance, the kitof the invention typically comprises directions for administration.

In the combination therapies of the invention, the compound of theinvention and the other therapeutic agent may be manufactured and/orformulated by the same or different manufacturers. Moreover, thecompound of the invention and the other therapeutic may be broughttogether into a combination therapy: (i) prior to release of thecombination product to physicians (e.g. in the case of a kit comprisingthe compound of the invention and the other therapeutic agent); (ii) bythe physician themselves (or under the guidance of the physician)shortly before administration; (iii) in the patient themselves, e.g.during sequential administration of the compound of the invention andthe other therapeutic agent. Accordingly, the invention provides the useof a compound of formula I′, I, II, Ill or IV, or a pharmaceuticallyacceptable salt thereof, for treating a disease or condition associatedwith neutral endopeptidase EC 3.4. 24.11 activity, wherein themedicament is prepared for administration with another therapeuticagent. The invention also provides the use of another therapeutic agentfor treating a disease or condition associated with neutralendopeptidase EC 3.4. 24.11 activity, wherein the medicament isadministered with a compound of formula I′, I, II, Ill or IV, or apharmaceutically acceptable salt thereof.

The invention also provides a compound of formula I′, I, III, III or IV,or a pharmaceutically acceptable salt thereof, for use in a method oftreating a disease or condition associated with neutral endopeptidase EC3.4. 24.11 activity, wherein the compound of formula I′, I, II, III orIV, or a pharmaceutically acceptable salt thereof, is prepared foradministration with another therapeutic agent. The invention alsoprovides another therapeutic agent for use in a method of treating adisease or condition associated with neutral endopeptidase EC 3.4. 24.11activity, wherein the other therapeutic agent is prepared foradministration with a compound of formula I′, I, II, III or IV, or apharmaceutically acceptable salt thereof. The invention also provides acompound of formula I′, I, II, III, or IV, or a pharmaceuticallyacceptable salt thereof, for use in a method of treating a disease orcondition associated with neutral endopeptidase EC 3.4. 24.11 activity,wherein the compound of formula I′, I, II, III or IV, or apharmaceutically acceptable salt thereof, is administered with anothertherapeutic agent. The invention also provides another therapeutic agentfor use in a method of treating a disease or condition associated withneutral endopeptidase EC 3.4. 24.11 activity, wherein the othertherapeutic agent is administered with a compound of formula I′, I, II,III or IV, or a pharmaceutically acceptable salt thereof.

The invention also provides the use of a compound of formula I′, I, II,III or IV, or a pharmaceutically acceptable salt thereof, for treating adisease or condition associated with neutral endopeptidase EC 3.4. 24.11activity, wherein the patient has previously (e.g. within 24 hours) beentreated with another therapeutic agent. The invention also provides theuse of another therapeutic agent for treating a disease or conditionassociated with neutral endopeptidase EC 3.4. 24.11 activity, whereinthe patient has previously (e.g. within 24 hours) been treated with acompound of formula I′, I, II, III or IV, or a pharmaceuticallyacceptable salt thereof.

In one embodiment, the other therapeutic agent is selected from:

In one embodiment, the other therapeutic agent is selected from:HMG-Co-A reductase inhibitor, an anigiotensin receptor blocker (ARBs,angiotensin II receptor antagonist), angiotensin converting enzyme (ACE)Inhibitor, a calcium channel blocker (CCB), an endothelin antagonist, arenin inhibitor, a diuretic, an ApoA-I mimic, an anti-diabetic agent, anobesity-reducing agent, an aldosterone receptor blocker, an endothelinreceptor blocker, an aldosterone synthase inhibitors (ASI), a CETPinhibitor and a phophodiesterase type 5 (PDE5) inhibitor.

The term “in combination with” a second agent or treatment includesco-administration of the compound of the invention (e.g., a compoundaccording to anyone of Formulae I-IV or a compound otherwise describedherein) with the second agent or treatment, administration of thecompound of the invention first, followed by the second agent ortreatment and administration of the second agent or treatment first,followed by the compound of the invention.

The term “second agent” includes any agent which is known in the art totreat, prevent, or reduce the symptoms of a disease or disorderdescribed herein, e.g. a disorder or disease responsive to theinhibition of neutral endopeptidase, such as for example, hypertension,resistant hypertension, pulmonary hypertension, pulmonary arterialhypertension, isolated systolic hypertension, peripheral vasculardisease, heart failure, congestive heart failure, left ventricularhypertrophy, angina, renal insufficiency (diabetic or non-diabetic),renal failure (including edema and salt retension), diabeticnephropathy, non-diabetic nephropathy, nephroic syndrome,glomerulonephritis, scleroderma, glomerular sclerosis, proteinurea ofprimary renal disease, renal vascular hypertention, diabetic retinopathyand end-stage renal disease (ESRD), endothelial dysfunction, diastolicdysfunction, hypertrophic cardiomyopathy, diabetic cardiac myopathy,supraventricular and ventricular arrhythmias, atrial fibrillation (AF),cardiac fibrosis, atrial flutter, detrimental vascular remodeling,plaque stabilization, myocardial infarction (MI), renal fibrosis,polycystic kidney disease (PKD), renal failure (including edema and saltretension), cyclical oedema, Menières disease, hyperaldosteroneism(primary and secondary) and hypercalciuria, ascites, glaucoma, menstrualdisorders, preterm labour, pre-eclampsia, endometriosis, andreproductive disorders (especially male and female infertility,polycystic ovarian syndrome, implantation failure), asthma, obstructivesleep apnea, inflammation, leukemia, pain, epilepsy, affective disorderssuch as depression and psychotic condition such as dementia andgeriatric confusion, obesity and gastrointestinal disorders (especiallydiarrhea and irritable bowel syndrome), wound healing (especiallydiabetic and venous ulcers and pressure sores), septic shock, themodulation of gastric acid secretion, the treatment of hyperreninaemia,cystic fibrosis, restenosis, type-2 diabetes, metabolic syndrome,diabetic complications and atherosclerosis, male and female sexualdysfunction.

Examples of second agents include HMG-Co-A reductase inhibitors,angiotensin II receptor antagonists, angiotensin converting enzyme (ACE)Inhibitors, calcium channel blockers (CCB), endothelin antagonists,renin inhibitors, diuretics, ApoA-I mimics, anti-diabetic agents,obesity-reducing agents, aldosterone receptor blockers, endothelinreceptor blockers, aldosterone synthase inhibitors (ASI) and CETPinhibitors.

The term “HMG-Co-A reductase inhibitor” (also calledbeta-hydroxy-beta-methylglutaryl-co-enzyme-A reductase inhibitors)includes active agents that may be used to lower the lipid levelsincluding cholesterol in blood. Examples include atorvastatin,cerivastatin, compactin, dalvastatin, dihydrocompactin, fluindostatin,fluvastatin, lovastatin, pitavastatin, mevastatin, pravastatin,rivastatin, simvastatin, and velostatin, or, pharmaceutically acceptablesalts thereof.

The term “ACE-inhibitor” (also called angiotensin converting enzymeinhibitors) includes molecules that interrupt the enzymatic degradationof angiotensin I to angiotensin II. Such compounds may be used for theregulation of blood pressure and for the treatment of congestive heartfailure. Examples include alacepril, benazepril, benazeprilat,captopril, ceronapril, cilazapril, delapril, enalapril, enaprilat,fosinopril, imidapril, lisinopril, moveltopril, perindopril, quinapril,ramipril, spirapril, temocapril, and trandolapril, or, pharmaceuticallyacceptables salt thereof.

The term “endothelin antagonist” includes bosentan (cf. EP 526708 A),tezosentan (cf. WO 96/19459), or, pharmaceutically acceptable saltsthereof.

The term “renin inhibitor” includes ditekiren (chemical name:[1S-[1R*,2R*,4R*(1R*,2R*)]]-1-[(1,1-dimethylethoxy)carbonyl]-L-prolyI-L-phenylalanyl-N-[2-hydroxy-5-methyl-1-(2-methylpropyl)-4-[[[(2-methyl-1-[[(2-pyridinylmethyl)amino]carbonyl]butyl]amino]carbonyl]hexyl]-N-alfa-methyl-L-histidinamide);terlakiren (chemical name:[R—(R*,S*)]—N-(4-morpholinylcarbonyl)-L-phenylalanyl-N-[1-(cyclohexylmethyl)-2-hydroxy-3-(1-methylethoxy)-3-oxopropyl]-S-methyl-L-cysteineamide);Aliskiren (chemical name:(2S,4S,5S,7S)-5-amino-N-(2-carbamoyl-2,2-dimethylethyl)-4-hydroxy-7-{[4-methoxy-3-(3-methoxypropoxy)phenyl]methyl}-8-methyl-2-(propan-2-yl)nonanamide) and zankiren(chemical name:[1S-[1R*[R*(R*)],2S*,3R]]—N-[1-(cyclohexylmethyl)-2,3-dihydroxy-5-methylhexyl]-alfa-[[2-[[(4-methyl-1-piperazinyl)sulfonyl]methyl]-1-oxo-3-phenylpropyl]-amino]-4-thiazolepropanamide),or, hydrochloride salts thereof, or, SPP630, SPP635 and SPP800 asdeveloped by Speedel, or RO 66-1132 and RO 66-1168 of Formula (A) and(B):

or, pharmaceutically acceptable salts thereof.

The term “aliskiren”, if not defined specifically, is to be understoodboth as the free base and as a salt thereof, especially apharmaceutically acceptable salt thereof, most preferably ahemi-fumarate salt thereof.

An angiotensin II receptor antagonist or a pharmaceutically acceptablesalt thereof is understood to be an active ingredient which bind to theAT₁-receptor subtype of angiotensin II receptor but do not result inactivation of the receptor. As a consequence of the inhibition of theAT₁ receptor, these antagonists can, for example, be employed asantihypertensives or for treating congestive heart failure.

The class of AT₁ receptor antagonists comprises compounds havingdiffering structural features, essentially preferred are thenon-peptidic ones. For example, mention may be made of the compoundswhich are selected from the group consisting of valsartan, losartan,candesartan, eprosartan, irbesartan, saprisartan, tasosartan,telmisartan, the compound with the designation E-1477 of the followingformula

the compound with the designation SC-52458 of the following formula

and the compound with the designation ZD-8731 of the following formula

or, in each case, a pharmaceutically acceptable salt thereof.

Preferred AT₁-receptor antagonist are those agents which have beenmarketed, most preferred is valsartan or a pharmaceutically acceptablesalt thereof.

The term “calcium channel blocker (CCB)” includes dihydropyridines(DHPs) and non-DHPs (e.g., diltiazem-type and verapamil-type CCBs).Examples include amlodipine, felodipine, ryosidine, isradipine,lacidipine, nicardipine, nifedipine, niguldipine, niludipine,nimodipine, nisoldipine, nitrendipine, and nivaldipine, and ispreferably a non-DHP representative selected from the group consistingof flunarizine, prenylamine, diltiazem, fendiline, gallopamil,mibefradil, anipamil, tiapamil and verapamil, or, pharmaceuticallyacceptable salts thereof. CCBs may be used as anti-hypertensive,anti-angina pectoris, or anti-arrhythmic drugs.

The term “diuretic” includes thiazide derivatives (e.g., chlorothiazide,hydrochlorothiazide, methylclothiazide, and chlorothalidon).

The term “ApoA-I mimic” includes D4F peptides (e.g., formulaD-W-F-K-A-F-Y-D-K-V-A-E-K-F-K-E-A-F)

The term “anti-diabetic agent” includes insulin secretion enhancers thatpromote the secretion of insulin from pancreatic β-cells. Examplesinclude biguanide derivatives (e.g., metformin), sulfonylureas (SU)(e.g., tolbutamide, chlorpropamide, tolazamide, acetohexamide,4-chloro-N-[(1-pyrrolidinylamino)carbonyl]-benzensulfonamide(glycopyramide), glibenclamide (glyburide), gliclazide,1-butyl-3-metanilylurea, carbutamide, glibonuride, glipizide,gliquidone, glisoxepid, glybuthiazole, glibuzole, glyhexamide,glymidine, glypinamide, phenbutamide, and tolylcyclamide), orpharmaceutically acceptable salts thereof. Further examples includephenylalanine derivatives (e.g., nateglinide[N-(trans-4-isopropylcyclohexylcarbonyl)-D-phenylalanine] (cf. EP 196222and EP 526171) of the formula

repaglinide[(S)-2-ethoxy-4-{2-[[3-methyl-1-[2-(1-piperidinyl)phenyl]butyl]amino]-2-oxoethyl}benzoicacid] (cf. EP 589874, EP 147850 A2, in particular Example 11 on page 61,and EP 207331 A1); calcium(2S)-2-benzyl-3-(cis-hexahydro-2-isoindolinlycarbonyl)-propionatedihydrate (e.g., mitiglinide (cf. EP 507534)); and glimepiride (cf. EP31058). Further examples include DPP-IV inhibitors, GLP-1 and GLP-1agonists.

DPP-IV is responsible for inactivating GLP-1. More particularly, DPP-IVgenerates a GLP-1 receptor antagonist and thereby shortens thephysiological response to GLP-1. GLP-1 is a major stimulator ofpancreatic insulin secretion and has direct beneficial effects onglucose disposal.

The DPP-IV inhibitor can be peptidic or, preferably, non-peptidic.DPP-IV inhibitors are in each case generically and specificallydisclosed e.g. in WO 98/19998, DE 196 16 486 A1, WO 00/34241 and WO95/15309, in each case in particular in the compound claims and thefinal products of the working examples, the subject-matter of the finalproducts, the pharmaceutical preparations and the claims are herebyincorporated into the present application by reference to thesepublications. Preferred are those compounds that are specificallydisclosed in Example 3 of WO 98/19998 and Example 1 of WO 00/34241,respectively.

GLP-1 is an insulinotropic protein which is described, e.g., by W. E.Schmidt et al. in Diabetologia, 28, 1985, 704-707 and in U.S. Pat. No.5,705,483.

The term “GLP-1 agonists” includes variants and analogs ofGLP-1(7-36)NH₂ which are disclosed in particular in U.S. Pat. No.5,120,712, U.S. Pat. No. 5,118,666, U.S. Pat. No. 5,512,549, WO 91/11457and by C. Orskov et al in J. Biol. Chem. 264 (1989) 12826. Furtherexamples include GLP-1(7-37), in which compound the carboxy-terminalamide functionality of Arg³⁶ is displaced with Gly at the 37^(th)position of the GLP-1(7-36)NH₂ molecule and variants and analogs thereofincluding GLN⁹-GLP-1(7-37), D-GLN⁹-GLP-1(7-37), acetyl LYS⁹-GLP-1(7-37),LYS¹⁸-GLP-1(7-37) and, in particular, GLP-1(7-37)OH, VAL⁸-GLP-1(7-37),GLY⁸-GLP-1(7-37), THR8-GLP-1(7-37), MET⁸-GLP-1(7-37) and4-imidazopropionyl-GLP-1. Special preference is also given to the GLPagonist analog exendin-4, described by Greig et al. in Diabetologia1999, 42, 45-50.

Also included in the definition “anti-diabetic agent” are insulinsensitivity enhancers which restore impaired insulin receptor functionto reduce insulin resistance and consequently enhance the insulinsensitivity. Examples include hypoglycemic thiazolidinedione derivatives(e.g., glitazone,(S)-((3,4-dihydro-2-(phenyl-methyl)-2H-1-benzopyran-6-yl)methyl-thiazolidine-2,4-dione(englitazone),5-{[4-(3-(5-methyl-2-phenyl-4-oxazolyl)-1-oxopropyl)-phenyl]-methyl}thiazolidine-2,4-dione(darglitazone),5-{[4-(1-methyl-cyclohexyl)methoxy)-phenyl]methyl}-thiazolidine-2,4-dione(ciglitazone),5-{[4-(2-(1-indolyl)ethoxy)phenyl]methyl}-thiazolidine-2,4-dione(DRF2189),5-{4-[2-(5-methyl-2-phenyl-4-oxazolyl)-ethoxy)]benzyl)thiazolidine-2,4-dione(BM-13.1246), 5-(2-naphthylsulfonyl)-thiazolidine-2,4-dione (AY-31637),bis{4-[(2,4-dioxo-5-thiazolidinyl)methyl]phenyl}methane (YM268),5-{4-[2-(5-methyl-2-phenyl-4-oxazolyl)-2-hydroxyethoxy]benzyl}-thiazolidine-2,4-dione(AD-5075),5-[4-(1-phenyl-1-cyclopropanecarbonylamino)-benzyl]-thiazolidine-2,4-dione(DN-108)5-{[4-(2-(2,3-dihydroindol-1-yl)ethoxy)phenyl]methyl}-thiazolidine-2,4-dione,5-[3-(4-chloro-phenyl])-2-propynyl]-5-phenylsulfonyl)thiazolidine-2,4-dione,5-[3-(4-chlorophenyl])-2-propynyl]-5-(4-fluorophenyl-sulfonyl)thiazolidine-2,4-dione,5-{[4-(2-(methyl-2-pyridinyl-amino)-ethoxy)phenyl]methyl}-thiazolidine-2,4-dione(rosiglitazone),5-{[4-(2-(5-ethyl-2-pyridyl)ethoxy)phenyl]-methyl}thiazolidine-2,4-dione(pioglitazone),5-{[4-((3,4-dihydro-6-hydroxy-2,5,7,8-tetramethyl-2H-1-benzopyran-2-yl)methoxy)-phenyl]-methyl}-thiazolidine-2,4-dione(troglitazone),5-[6-(2-fluoro-benzyloxy)naphthalen-2-ylmethyl]-thiazolidine-2,4-dione(MCC555),5-{[2-(2-naphthyl)-benzoxazol-5-yl]-methyl}thiazolidine-2,4-dione(T-174) and5-(2,4-dioxothiazolidin-5-ylmethyl)-2-methoxy-N-(4-trifluoromethyl-benzyl)benzamide(KRP297)).

Further anti-diabetic agents include, insulin signalling pathwaymodulators, like inhibitors of protein tyrosine phosphatases (PTPases),antidiabetic non-small molecule mimetic compounds and inhibitors ofglutamine-fructose-6-phosphate amidotransferase (GFAT); compoundsinfluencing a dysregulated hepatic glucose production, like inhibitorsof glucose-6-phosphatase (G6 Pase), inhibitors offructose-1,6-bisphosphatase (F-1,6-Bpase), inhibitors of glycogenphosphorylase (GP), glucagon receptor antagonists and inhibitors ofphosphoenolpyruvate carboxykinase (PEPCK); pyruvate dehydrogenase kinase(PDHK) inhibitors; inhibitors of gastric emptying; insulin; inhibitorsof GSK-3; retinoid X receptor (RXR) agonists; agonists of Beta-3 AR;agonists of uncoupling proteins (UCPs); non-glitazone type PPARγagonists; dual PPARα/PPARγ agonists; antidiabetic vanadium containingcompounds; incretin hormones, like glucagon-like peptide-1 (GLP-1) andGLP-1 agonists; beta-cell imidazoline receptor antagonists; miglitol;α₂-adrenergic antagonists; and pharmaceutically acceptable saltsthereof.

The term “obesity-reducing agent” includes lipase inhibitors (e.g.,orlistat) and appetite suppressants (e.g., sibutramine and phentermine).

An aldosterone synthase inhibitor or a pharmaceutically acceptable saltthereof is understood to be an active ingredient that has the propertyto inhibit the production of aldosterone. Aldosterone synthase (CYP11B2)is a mitochondrial cytochrome P450 enzyme catalyzing the last step ofaldosterone production in the adrenal cortex, i.e., the conversion of11-deoxycorticosterone to aldosterone. The inhibition of the aldosteroneproduction with so-called aldosterone synthase inhibitors is known to bea successful variant to treatment of hypokalemia, hypertension,congestive heart failure, atrial fibrillation or renal failure. Suchaldosterone synthase inhibition activity is readily determined by thoseskilled in the art according to standard assays (e.g., US 2007/0049616).

The class of aldosterone synthase inhibitors comprises both steroidaland non-steroidal aldosterone synthase inhibitors, the later being mostpreferred.

Preference is given to commercially available aldosterone synthaseinhibitors or those aldosterone synthase inhibitors that have beenapproved by the health authorities.

The class of aldosterone synthase inhibitors comprises compounds havingdiffering structural features. For example, mention may be made of thecompounds which are selected from the group consisting of thenon-steroidal aromatase inhibitors anastrozole, fadrozole (including the(+)-enantiomer thereof), as well as the steroidal aromatase inhibitorexemestane, or, in each case where applicable, a pharmaceuticallyacceptable salt thereof.

The most preferred non-steroidal aldosterone synthase inhibitor is the(+)-enantiomer of the hydrochloride of fadrozole (U.S. Pat. Nos.4,617,307 and 4,889,861) of formula

or, if appropriable, a pharmaceutically acceptable salt thereof.

A preferred steroidal aldosterone antagonist is eplerenone (cf. EP122232 A) of the formula

or Spironolactone; or, in each case, if appropriable, a pharmaceuticallyacceptable salt thereof.

Aldosterone synthase inhibitors useful in said combination are compoundsand analogs generically and specifically disclosed e.g. inUS2007/0049616, in particular in the compound claims and the finalproducts of the working examples, the subject-matter of the finalproducts, the pharmaceutical preparations and the claims are herebyincorporated into the present application by reference to thispublication. Preferred aldosterone synthase inhibitors suitable for usein the present invention include, without limitation4-(6,7-dihydro-5H-pyrrolo[1,2-c]imidazol-5-yl)-3-methylbenzonitrile;5-(2-chloro-4-cyanophenyl)-6,7-dihydro-5H-pyrrolo[1,2-c]imidazole-5-carboxylicacid (4-methoxybenzyl)methylamide;4′-fluoro-6-(6,7,8,9-tetrahydro-5H-imidazo[1,5-a]azepin-5-yl)biphenyl-3-carbonitrile;5-(4-Cyano-2-methoxyphenyl)-6,7-dihydro-5H-pyrrolo[1,2-c]imidazole-5-carboxylicacid butyl ester;4-(6,7-Dihydro-5H-pyrrolo[1,2-c]imidazol-5-yl)-2-methoxybenzonitrile;5-(2-Chloro-4-cyanophenyl)-6,7-dihydro-5H-pyrrolo[1,2-c]imidazole-5-carboxylicacid 4-fluorobenzyl ester;5-(4-Cyano-2-trifluoromethoxyphenyl)-6,7-dihydro-5H-pyrrolo[1,2-c]imidazole-5-carboxylicacid methyl ester;5-(4-Cyano-2-methoxyphenyl)-6,7-dihydro-5H-pyrrolo[1,2-c]imidazole-5-carboxylicacid 2-isopropoxyethyl ester;4-(6,7-Dihydro-5H-pyrrolo[1,2-c]imidazol-5-yl)-2-methylbenzonitrile;4-(6,7-dihydro-5H-pyrrolo[1,2-c]imidazol-5-yl)-3-fluorobenzonitrile;4-(6,7-Dihydro-5H-pyrrolo[1,2-c]imidazol-5-yl)-2-methoxybenzonitrile;3-Fluoro-4-(7-methylene-6,7-dihydro-5H-pyrrolo[1,2-d]imidazol-5-yl)benzonitrile;cis-3-Fluoro-4-[7-(4-fluoro-benzyl)-5,6,7,8-tetrahydro-imidazo[1,5-a]pyridin-5-yl]benzonitrile;4′-Fluoro-6-(9-methyl-6,7,8,9-tetrahydro-5H-imidazo[1,5-a]azepin-5-yl)biphenyl-3-carbonitrile;4′-Fluoro-6-(9-methyl-6,7,8,9-tetrahydro-5H-imidazo[1,5-a]azepin-5-yl)biphenyl-3-carbonitrileor in each case, the (R) or (S) enantiomer thereof; or if appropriable,a pharmaceutically acceptable salt thereof.

The term aldosterone synthase inhibitors also include compounds andanalogs disclosed in WO2008/076860, WO2008/076336, WO2008/076862,WO2008/027284, WO2004/046145, WO2004/014914, WO2001/076574.

Furthermore Aldosterone synthase inhibitors also include compounds andanalogs disclosed in U.S. patent applications US2007/0225232,US2007/0208035, US2008/0318978, US2008/0076794, US2009/0012068,US20090048241 and in PCT applications WO2006/005726, WO2006/128853,WO2006128851, WO2006/128852, WO2007065942, WO2007/116099, WO2007/116908,WO2008/119744 and in European patent application EP 1886695. Preferredaldosterone synthase inhibitors suitable for use in the presentinvention include, without limitation8-(4-Fluorophenyl)-5,6-dihydro-8H-imidazo[5,1-c][1,4]oxazine;4-(5,6-Dihydro-8H-imidazo[5,1-c][1,4]oxazin-8-yl)-2-fluorobenzonitrile;4-(5,6-Dihydro-8H-imidazo[5,1-c][1,4]oxazin-8-yl)-2,6-difluorobenzonitrile;4-(5,6-Dihydro-8H-imidazo[5,1-c][1,4]oxazin-8-yl)-2-methoxybenzonitrile;3-(5,6-Dihydro-8H-imidazo[5,1-c][1,4]oxazin-8-yl)benzonitrile;4-(5,6-Dihydro-8H-imidazo[5,1-c][1,4]oxazin-8-yl)phthalonitrile;4-(8-(4-Cyanophenyl)-5,6-dihydro-8H-imidazo[5,1-c][1,4]oxazin-8-yl)benzonitrile;4-(5,6-Dihydro-8H-imidazo[5,1-c][1,4]oxazin-8-yl)benzonitrile;4-(5,6-Dihydro-8H-imidazo[5,1-c][1,4]oxazin-8-yl)naphthalene-1-carbonitrile;8-[4-(1H-Tetrazol-5-yl)phenyl]-5,6-dihydro-8H-imidazo[5,1-c][1,4]oxazineas developed by Speedel or in each case, the (R) or (S) enantiomerthereof; or if appropriable, a pharmaceutically acceptable salt thereof.

The term “endothelin receptor blocker” includes bosentan. The term “CETPinhibitor” refers to a compound that inhibits the cholesteryl estertransfer protein (CETP) mediated transport of various cholesteryl estersand triglycerides from HDL to LDL and VLDL. Such CETP inhibitionactivity is readily determined by those skilled in the art according tostandard assays (e.g., U.S. Pat. No. 6,140,343). Examples includecompounds disclosed in U.S. Pat. No. 6,140,343 and U.S. Pat. No.6,197,786 (e.g.,[2R,4S]4-[(3,5-bis-trifluoromethyl-benzyl)-methoxycarbonyl-amino]-2-ethyl-6-trifluoromethyl-3,4-dihydro-2H-quinoline-1-carboxylicacid ethyl ester (torcetrapib); compounds disclosed in U.S. Pat. No.6,723,752 (e.g.,(2R)-3-{[3-(4-Chloro-3-ethyl-phenoxy)-phenyl]-[[3-(1,1,2,2-tetrafluoro-ethoxy)-phenyl]-methyl]-amino}-1,1,1-trifluoro-2-propanol);compounds disclosed in U.S. patent application Ser. No. 10/807,838;polypeptide derivatives disclosed in U.S. Pat. No. 5,512,548;rosenonolactone derivatives and phosphate-containing analogs ofcholesteryl ester disclosed in J. Antibiot., 49(8): 815-816 (1996), andBioorg. Med. Chem. Lett.; 6:1951-1954 (1996), respectively. Furthermore,the CETP inhibitors also include those disclosed in WO2000/017165,WO2005/095409 and WO2005/097806.

A preferred PDE5 inhibitor is Sildenafil.

Second agent of particular interest include Endothelin antagonists,renin inhibitors, angiotensin II receptor antagonists, calcium channelblockers, diuretics, antidiabetic agents such as DPPIV inhibitors, andaldosterone synthase inhibitors.

In one embodiment, the invention provides a combination, in particular apharmaceutical combination, comprising a therapeutically effectiveamount of the compound according to the definition of formula I′, II,III or IV or a pharmaceutically acceptable salt thereof, and one or moretherapeutically active agents selected from HMG-Co-A reductaseinhibitors, angiotensin II receptor antagonists, angiotensin convertingenzyme (ACE) Inhibitors, calcium channel blockers (CCB), endothelinantagonists, renin inhibitors, diuretics, ApoA-I mimics, anti-diabeticagents, obesity-reducing agents, aldosterone receptor blockers,endothelin receptor blockers, aldosterone synthase inhibitors (ASI) andCETP inhibitors.

In one embodiment, the invention provides a method of inhibiting neutralendopeptidase EC 3.4. 24.11 activity in a subject, wherein the methodcomprises administering to the subject a therapeutically effectiveamount of the compound according to the definition of formula I′, II,III or IV or a pharmaceutically acceptable salt thereof.

In one embodiment, the invention provides a method of treating adisorder or a disease in a subject associated with neutral endopeptidaseEC 3.4. 24.11 activity, wherein the method comprises administering tothe subject a therapeutically effective amount of the compound accordingto the definition of formula I′, I, II, III or IV or a pharmaceuticallyacceptable salt thereof.

In one embodiment, the invention provides a method of treating adisorder or a disease in a subject associated with neutral endopeptidaseEC 3.4. 24.11 activity, wherein the disorder or the disease is selectedfrom hypertension, resistant hypertension, pulmonary hypertension,pulmonary arterial hypertension, isolated systolic hypertension,peripheral vascular disease, heart failure, congestive heart failure,left ventricular hypertrophy, angina, renal insufficiency (diabetic ornon-diabetic), renal failure (including edema and salt retension),diabetic nephropathy, nephroic syndrome, glomerulonephritis,scleroderma, glomerular sclerosis, proteinurea of primary renal disease,renal vascular hypertention, diabetic retinopathy and end-stage renaldisease (ESRD), endothelial dysfunction, diastolic dysfunction,hypertrophic cardiomyopathy, diabetic cardiac myopathy, supraventricularand ventricular arrhythmias, atrial fibrillation (AF), cardiac fibrosis,atrial flutter, detrimental vascular remodeling, plaque stabilization,myocardial infarction (MI), renal fibrosis, polycystic kidney disease(PKD), renal failure (including edema and salt retension), cyclicaloedema, Menières disease, hyperaldosteroneism (primary and secondary)and hypercalciuria, ascites, glaucoma, menstrual disorders, pretermlabour, pre-eclampsia, endometriosis, and reproductive disorders(especially male and female infertility, polycystic ovarian syndrome,implantation failure), asthma, obstructive sleep apnea, inflammation,leukemia, pain, epilepsy, affective disorders such as depression andpsychotic condition such as dementia and geriatric confusion, obesityand gastrointestinal disorders (especially diarrhea and irritable bowelsyndrome), wound healing (especially diabetic and venous ulcers andpressure sores), septic shock, gastric acid secretion dysfunction,hyperreninaemia, cystic fibrosis, restenosis, type-2 diabetes, metabolicsyndrome, diabetic complications and atherosclerosis, male and femalesexual dysfunction.

In one embodiment, the invention provides a compound according to thedefinition of formula I′, I, II, III or IV, for use as a medicament.

In one embodiment, the invention provides the use of a compoundaccording to the definition of formula I′, I, II, Ill or IV or apharmaceutically acceptable salt thereof, for the treatment of adisorder or disease in a subject associated with neutral endopeptidaseEC 3.4. 24.11 activity.

In one embodiment, the invention provides the use of a compoundaccording to the definition of formula I′, I, II, Ill or IV, in themanufacture of a medicament for the treatment of a disorder or diseasein a subject characterized by an activity of neutral endopeptidase EC3.4. 24.11, wherein said disorder or disease is in particular selectedfrom hypertension, resistant hypertension, pulmonary hypertension,pulmonary arterial hypertension, isolated systolic hypertension,peripheral vascular disease, heart failure, congestive heart failure,left ventricular hypertrophy, angina, renal insufficiency (diabetic ornon-diabetic), renal failure (including edema and salt retension),diabetic nephropathy, non-diabetic nephropathy, nephroic syndrome,glomerulonephritis, scleroderma, glomerular sclerosis, proteinurea ofprimary renal disease, renal vascular hypertention, diabetic retinopathyand end-stage renal disease (ESRD), endothelial dysfunction, diastolicdysfunction, hypertrophic cardiomyopathy, diabetic cardiac myopathy,supraventricular and ventricular arrhythmias, atrial fibrillation (AF),cardiac fibrosis, atrial flutter, detrimental vascular remodeling,plaque stabilization, myocardial infarction (MI), renal fibrosis,polycystic kidney disease (PKD), renal failure (including edema and saltretension), cyclical oedema, Menières disease, hyperaldosteroneism(primary and secondary) and hypercalciuria, ascites, glaucoma, menstrualdisorders, preterm labour, pre-eclampsia, endometriosis, andreproductive disorders (especially male and female infertility,polycystic ovarian syndrome, implantation failure), asthma, obstructivesleep apnea, inflammation, leukemia, pain, epilepsy, affective disorderssuch as depression and psychotic condition such as dementia andgeriatric confusion, obesity and gastrointestinal disorders (especiallydiarrhea and irritable bowel syndrome), wound healing (especiallydiabetic and venous ulcers and pressure sores), septic shock, gastricacid secretion dysfunction, hyperreninaemia, cystic fibrosis,restenosis, type-2 diabetes, metabolic syndrome, diabetic complicationsand atherosclerosis, male and female sexual dysfunction

In one embodiment, the invention provides the use of a compoundaccording to the definition of formula I′, I, II, III or IV, for thetreatment of a disorder or disease in a subject characterized by anactivity of neutral endopeptidase EC 3.4. 24.11, wherein the disorder ordisease is selected from hypertension, resistant hypertension, pulmonaryhypertension, pulmonary arterial hypertension, isolated systolichypertension, peripheral vascular disease, heart failure, congestiveheart failure, left ventricular hypertrophy, angina, renal insufficiency(diabetic or non-diabetic), renal failure (including edema and saltretension), diabetic nephropathy, non-diabetic nephropathy, nephroicsyndrome, glomerulonephritis, scleroderma, glomerular sclerosis,proteinurea of primary renal disease, renal vascular hypertention,diabetic retinopathy and end-stage renal disease (ESRD), endothelialdysfunction, diastolic dysfunction, hypertrophic cardiomyopathy,diabetic cardiac myopathy, supraventricular and ventricular arrhythmias,atrial fibrillation (AF), cardiac fibrosis, atrial flutter, detrimentalvascular remodeling, plaque stabilization, myocardial infarction (MI),renal fibrosis, polycystic kidney disease (PKD), renal failure(including edema and salt retension), cyclical oedema, Menières disease,hyperaldosteroneism (primary and secondary) and hypercalciuria, ascites,glaucoma, menstrual disorders, preterm labour, pre-eclampsia,endometriosis, and reproductive disorders (especially male and femaleinfertility, polycystic ovarian syndrome, implantation failure), asthma,obstructive sleep apnea, inflammation, leukemia, pain, epilepsy,affective disorders such as depression and psychotic condition such asdementia and geriatric confusion, obesity and gastrointestinal disorders(especially diarrhea and irritable bowel syndrome), wound healing(especially diabetic and venous ulcers and pressure sores), septicshock, gastric acid secretion dysfunction, hyperreninaemia, cysticfibrosis, restenosis, type-2 diabetes, metabolic syndrome, diabeticcomplications and atherosclerosis, male and female sexual dysfunction

EXEMPLIFICATION OF THE INVENTION

The following examples are intended to illustrate the invention and arenot to be construed as being limitations thereon. Temperatures are givenin degrees centigrade. If not mentioned otherwise, all evaporations areperformed under reduced pressure, typically between about 15 mm Hg and100 mm Hg (=20-133 mbar). The structure of final products, intermediatesand starting materials is confirmed by standard analytical methods,e.g., microanalysis and spectroscopic characteristics, e.g., MS, IR,NMR. Abbreviations used are those conventional in the art.

All starting materials, building blocks, reagents, acids, bases,dehydrating agents, solvents, and catalysts utilized to synthesis thecompounds of the present invention are either commercially available orcan be produced by organic synthesis methods known to one of ordinaryskill in the art (Houben-Weyl 4th Ed. 1952, Methods of OrganicSynthesis, Thieme, Volume 21). Further, the compounds of the presentinvention can be produced by organic synthesis methods known to one ofordinary skill in the art as shown in the following examples.

Exemplification of the Invention Abbreviations

br: broad bs: broad singlet Ac: Acetyl Atm: atmosphere Aq: aqueouscalcd: calculated Bn: benzyl Boc: tert-butoxycarbonyl d: doublet dd:doublet of doublets DCM: dichloromethane DME: 1,4-dimethoxyethane DMF:N,N-dimethylformamide DMSO: dimethylsulfoxide DAD: diode array detectorDTT: dithiothreitol EDTA: ethylenediamine tetraacetic acid ESI:electrospray ionization Et and EtOAc: ethyl and ethyl acetate HATU:O-(7-azobenzotriazol-1-yl)- HOBt: 1-hydroxy-7- 1,1,3,3- azabenzotriazoletetramethyluroniumhexafluorophosphate HPLC: high pressure liquid LC andLCMS: liquid chromatography chromatography and liquid HPLC-RT: highpressure liquid chromatography and mass chromatography-retention timespectrometry H: Hour(s) IR: infrared MeOH: methanol MS: massspectrometry m: multiplet min: minutes Me: methyl m/z: mass to chargeratio M and mM: Molar and millimole(s) Mg: milligram n.d.: notdetermined NMR: nuclear magnetic resonance PMBCl: para-methoxybenzylchloride Pr and iPr: propyl and isopropyl ppm: parts per million Pd/C:Palladium on Carbon Ph: Phenyl q: quartet RP: reverse phase RT: roomtemperature s: singlet t: triplet TFA: trifluoroacetic acid THE:tetrahydrofuran TLC: thin layer chromatography tBu: tert-butyl μL, mLand L: microlitre, millilitre Tris•HCl: and litreaminotris(hydroxymethyl) methane hydrochloride wt: weight UV:ultraviolet

The following examples are intended to illustrate the invention and arenot to be construed as being limitations thereon. Temperatures are givenin degrees centigrade. If not mentioned otherwise, all evaporations areperformed under reduced pressure, preferably between about 15 mm Hg and100 mm Hg (=20-133 mbar). The structure of final products, intermediatesand starting materials is confirmed by standard analytical methods,e.g., microanalysis and spectroscopic characteristics, e.g., MS, IR,NMR. Abbreviations used are those conventional in the art.

All starting materials, building blocks, reagents, acids, bases,dehydrating agents, solvents, and catalysts utilized to synthesis thecompounds of the present invention are either commercially available orcan be produced by organic synthesis methods known to one of ordinaryskill in the art (Houben-Weyl 4th Ed. 1952, Methods of OrganicSynthesis, Thieme, Volume 21). Further, the compounds of the presentinvention can be produced by organic synthesis methods known to one ofordinary skill in the art as shown in the following examples.

The compounds in the examples 3-1 and 3-2 to 3-15 have been found tohave IC₅₀ values in the range of about 0.001 nM to about 10,000 nM forNEP.

The conditions for measuring the retention times are as follows:

HPLC Condition A:

Column: INERTSIL C8-3, 3 μm×33 mm×3.0 mm at 40° C.Flow rate: 2 mL/minMobile phase: A) 0.5 mM ammonium formate in H₂O; B) 50% MeOH in CH₃CNGradient: linear gradient from 5% B to 95% B in 2 min

Detection: DAD-UV at 210-400 nm HPLC Condition B:

Column: INERTSIL C8-3, 3 μm×33 mm×3.0 mm at 40° C.

Flow rate: 2 mL/minMobile phase: A) 0.1% formic acid in H₂O; B) 50% MeOH in CH₃CNGradient: linear gradient from 5% B to 95% B in 2 min

Detection: DAD-UV at 210-400 nm HPLC Condition C:

Column: INERTSIL C8-3, 3 μm×33 mm×3.0 mm at 40° C.

Flow rate: 2 mL/minMobile phase: A) 0.5 mM ammonium formate in H₂O; B) 50% MeOH in CH₃CNGradient: linear gradient from 40% B to 95% B in 2 min

Detection: DAD-UV at 210-400 nm HPLC Condition D:

Column: INERTSIL C8-3, 3 μm×33 mm×3.0 mm at 40° C.

Flow rate: 2 mL/minMobile phase: A) 0.1% formic acid in H₂O; B) 50% MeOH in CH₃CNGradient: linear gradient from 40% B to 95% B in 2 min

Detection: DAD-UV at 210-400 nm

The relative stereochemistry was determined using two dimensional NMR.Under the reaction conditions, the racemization of the stereocenterbearing the bis-phenylmethyl group is not expected. Therefore theabsolute stereochemistry was determined based on the relativestereochemistry and the stereochemistry of the center bearing thebisphenylmethyl group.

Example 1-1 Synthesis of(S)-2-[(S)-2-(3′-chloro-biphenyl-4-yl)-1-(1H-tetrazol-5-ylcarbamoyl)-ethylamino]-propionicacid ethyl ester

To a suspension of(S)-3-(3′-chloro-biphenyl-4-yl)-2-((S)-1-ethoxycarbonyl-ethylamino)-propionicacid (4.0 g, 10.84 mmol) in dichloromethane (60 mL) and saturatedaqueous NaHCO₃ (10 mL) was added triphosgene (1.90 g, 6.39 mmol). Aftervigorously stirred for 0.5 hour, the reaction mixture was diluted withEtOAc and partially concentrated under reduced pressure. Excess oftriphosgene was quenched by adding saturated aqueous NaHCO₃ and stirredfor 0.5 hour. The mixture was extracted with EtOAc and washed withbrine. The organic layer was dried over Na₂SO₄ and concentrated underreduced pressure. The obtained residue was dissolved in dichloromethane(50 mL). To the mixture was added triethylamine (1.93 mL, 13.8 mmol) and5-amino-1H-tetrazole (1.18 g, 13.84 mmol) at 0° C., and the reactionmixture was gradually warmed to room temperature. After stirred for 2hours, the reaction mixture was concentrated and purified by silica gelcolumn chromatography (eluent: 10% MeOH in dichloromethane) to give amixture of the desired trans isomer product and the cis isomer. Theobtained material was re-crystallized from CH₃CN three times to give(S)-2-[(S)-2-(3′-chloro-biphenyl-4-yl)-1-(1H-tetrazol-5-ylcarbamoyl)-ethylamino]-propionicacid ethyl ester (3.92 g). ¹H NMR (400 MHz, DMSO-d6) δ 1.11 (t, 3H,J=7.1 Hz), 1.15 (d, 3H, J=6.8 Hz), 2.89 (dd, 1H, J=8.1, 13.7 Hz), 3.02(dd, 1H, J=5.8, 14.0 Hz), 3.27-3.36 (m, 1H), 3.75-3.83 (m, 1H), 4.01 (q,2H, J=7.1 Hz), 7.34 (d, 2H, J=8.3 Hz), 7.38-7.42 (m, 1H), 7.47 (dd, 1H,J=7.8, 7.8 Hz), 7.60-7.65 (m, 3H), 7.69 (dd, 1H, J=1.8, 1.8 Hz); MS: m/z(MH⁺) 443; HRMS: calculated for C₂₂H₂₃ClN₆O₃ (M)⁺442.1. found 442.1; EA:Calculated for C21H23ClN6O3: C, 56.95; H, 5.23; N, 18.97. Found: C,56.88; H, 5.07; N, 19.1.

Chiral HPLC retention time=9.26 min. [condition: Daicel CHIRALCEL OJ-H4.6×100 mm); flow rate=1 ml/min.; eluent: 20% EtOH (with 0.1% TFA) inheptane].

Following compounds were prepared using similar procedure as example 1-1with appropriate intermediates:

Ex- HPLC- am- RT MS ple (con- (M + # Product Intermediates dition) 1)Ex- am- ple 1-2

  (S)-2-[(S)-2-(2′,5′- Dichloro-biphenyl- 4-yl)-1-(1H-tetrazol-5-ylcarbamoyl)- ethylamino]- propionic acid tert- butyl ester

  (S)-2-((S)-1-tert- Butoxycarbonyl- ethylamino)-3-(2′,5′-dichloro-biphenyl- 4-yl)-propionic acid And  

  5-amino-1H- tetrazole 1.38 min (C) 505 Ex- am- ple 1-3

  (S)-2-[(S)-2-Biphenyl-4- yl-1-(1H-tetrazol-5- ylcarbamoyl)-ethylamino]-propionic acid ethyl ester

  (S)-3-Biphenyl-4-yl- 2-((S)-1-ethoxycarbonyl- ethylamino)- propionicacid And  

  5-amino-1H- tetrazole 1.55 min (A) 409 Ex- am- ple 1-4

  (S)-2-[(S)-2-(5′-Fluoro- 2′-methoxy- biphenyl-4-yl)-1-(1H-tetrazol-5-ylcarbamoyl)- ethylamino]- propionic acid tert-butyl ester

  (S)-2-((S)-1-tert- Butoxycarbonyl- ethylamino)-3-(5′-fluoro-2′-methoxy- biphenyl-4-yl)- propionic acid And  

  5-amino-1H- tetrazole 1.27 min (C) 485 Ex- am- ple 1-5

  5-[(S)-3-Biphenyl-4-yl- 2-((S)-1-tert- butoxycarbonyl- ethylamino)-propionylamino]-1H- pyrazole-3-carboxylic acid methyl ester

  (S)-3-Biphenyl-4-yl- 2-((S)-1-tert- butoxycarbonyl- ethylamino)-propionic acid And  

  5-amino-1H- pyrazole-3- carboxylic acid methyl ester 1.49 min (C) 493Ex- am- ple 1-6

  (S)-2-[(S)-2-(3′-Chloro- biphenyl-4- yl)-1-(3-hydroxy-isoxazol-5-ylcarbamoyl)- ethylamino]- propionic acid tert- butyl ester

  (S)-2-((S)-1-tert- Butoxycarbonyl- ethylamino)-3-(3′-chloro-biphenyl-4- yl)-propionic acid And  

  5-amino-isoxazol-3-ol 1.48 min (C) 486 Ex- am- ple 1-7

  (S)-2-[(S)-2-(3′-Chloro- biphenyl-4-yl)-1-(1- methyl-1H-tetrazol-5-ylcarbamoyl)- ethylamino]-propionic acid ethyl ester

  (S)-3-(3′-Chloro- biphenyl-4-yl)-2-((S)- 1-ethoxycarbonyl-ethylamino)- propionic acid And  

  1-methyl-1H- tetrazol-5-ylamine 1.12 min (C) 457 Ex- am- ple 1-8

  (S)-2-[(S)-2-(3′-Chloro- biphenyl-4-yl)-1-(1H-tetrazol-5-ylcarbamoyl)- ethylamino]-4-phenyl- butyric acid ethyl ester

  (S)-2-[(S)-1-carboxy- 2-(3′-chloro- biphenyl-4-yl)- ethylamino]-4-phenyl-butyric acid ethyl ester And  

  5-amino-1H- tetrazole 1.47 min (C) 533.4 Ex- am- ple 1-9

  (S)-2-[(S)-2-(3′-Chloro- biphenyl-4-yl)-1-(1H-tetrazol-5-ylcarbamoyl)- ethylamino]-butyric acid tert-butyl ester

  (S)-2-[(S)-1- Carboxy-2-(3′- chloro-biphenyl-4- yl)-ethylamino]-butyric acid tert-butyl ester And  

  5-amino-1H- tetrazole 1.27 (C) 485 Ex- am- ple 1-10

  2-[2-(3′-Chloro- biphenyl-4-yl)-1-(1H- tetrazol-5-ylcarbamoyl)-ethylamino]-3-phenyl- propionic acid ethyl ester

  (S)-3-(3′-Chloro- biphenyl-4-yl)-2- ((S)-1- ethoxycarbonyl-2-phenyl-ethyl amino)-propionic acid And  

  5-amino-1H- tetrazole 1.31 (C) 519

Experiment 1-11(S)-2-{(S)-2-(3′-chloro-biphenyl-4-yl)-1-[methyl-(1H-tetrazol-5-yl)-carbamoyl]-ethylamino}-propionicacid ethyl ester

To a suspension of(S)-3-(3′-chloro-biphenyl-4-yl)-2-((S)-1-ethoxycarbonyl-ethylamino)-propionicacid (225 mg, 0.599 mmol) in dichloromethane (4 mL) and saturatedaqueous NaHCO₃ (1 mL) was added triphosgene (178 mg, 0.599 mmol). Aftervigorously stirred for 10 min, the reaction mixture was diluted withEtOAc and partially concentrated under reduced pressure. Excess oftriphosgene was quenched by adding saturated aqueous NaHCO₃ and stirredfor 0.5 hour. The mixture was extracted with EtOAc and washed withbrine. The organic layer was dried over Na₂SO₄ and concentrated underreduced pressure. The obtained residue was dissolved in dichloromethane(5 mL). To the mixture were added triethylamine (0.167 mL, 1.197 mmol)and [1-(4-methoxy-benzyl)-1H-tetrazol-5-yl]-methyl-amine (197 mg, 0.898mmol) and stirred at 45° C. overnight. Additional triethylamine (0.167mL, 1.197 mmol) and [1-(4-methoxy-benzyl)-1H-tetrazol-5-yl]-methyl-amine(197 mg, 0.898 mmol) were added and stirred at 45° C. for 30 hours. Thereaction mixture was concentrated under reduced pressure and purified bysilica gel column chromatography (eluent: 10% MeOH in DCM) to give(S)-2-((S)-2-(3′-chloro-biphenyl-4-yl)-1-{[1-(4-methoxy-benzyl)-1H-tetrazol-5-yl]-methyl-carbamoyl}ethylamino)-propionicacid ethyl ester (261 mg). MS: m/z (MW) 577; HPLC retention time 1.36min (HPLC condition C).

Next,(S)-2-((S)-2-(3′-chloro-biphenyl-4-yl)-1-{[1-(4-methoxy-benzyl)-1H-tetrazol-5-yl]-methyl-carbamoyl}-ethylamino)-propionicacid ethyl ester (260 mg, 0.451 mmol) was dissolved in TFA (5 mL) andDCM (5 mL) and stirred at 50° C. for 12 hours and at 75° C. for 5 hours.The reaction mixture was concentrated under reduced pressure to give(S)-2-{(S)-2-(3′-chloro-biphenyl-4-yl)-1-[methyl-(1H-tetrazol-5-yl)-carbamoyl]-ethylamino}-propionicacid ethyl ester (120 mg). MS: m/z (MW) 457; HPLC retention time 0.95min (HPLC condition C).

Following compounds were prepared using similar procedure as example 1-1with appropriate intermediates:

HPLC-RT MS Example # Product Intermediates (condition) (M + 1) Example1-12

  (S)-3-benzyloxy-2-[(S)- 2-(3′-chloro-biphenyl-4- yl)-1-(1H-tetrazol-5-ylcarbamoyl)- ethylamino]-propionic acid ethyl ester

  (S)-2-((S)-2- benzyloxy-1- ethoxycarbonyl- ethylamino)-3-(3′-chloro-biphenyl-4- yl)-propionic acid And  

  5-amino-1H- tetrazole 1.31 min (C) 549 Example 1-13

  (S)-2-[(S)-2-(3′-chloro- biphenyl-4-yl)-1-(1H-tetrazol-5-ylcarbamoyl)- ethylamino]-3-methoxy- propionic acid ethylester

  (S)-3-(3′-chloro- biphenyl-4-yl)-2-((S)- 1-ethoxycarbonyl-2- methoxy-ethylamino)- propionic acid And  

  5-amino-1H- tetrazole 1.29 min (A) 471

Example 1-14(S)-2-[(S)-2-(3′-chloro-biphenyl-4-yl)-1-(1H-tetrazol-5-ylcarbamoyl)-ethoxy]-propionicacid ethyl ester

To a solution of(S)-3-(3′-chloro-biphenyl-4-yl)-2-((S)-1-ethoxycarbonyl-ethoxy)-propionicacid (62 mg, 0.165 mmol) in THF (5 ml) at room temperature was added5-aminotetrazole (38.0 mg, 0.447 mmol), DIPEA (0.086 ml, 0.494 mmol) andfollowed by 1,3-diisopropylcarbodiimide (0.060 ml, 0.387 mmol). Thereaction was stirred at room temperature for 3 hr. The reaction wasquenched by brine and was extracted with EtOAc. The combined organiclayer was washed with brine and dried over anhydrous sodium sulfate,filtered and concentrated. HPLC retention time=0.99 minutes (conditionC); MS (m+1)=444.

Example 2-1(S)-2-[(R)-2-(3′-chloro-biphenyl-4-yl)-1-(1H-tetrazol-5-ylcarbamoyl)-ethylamino]-propionicacid ethyl ester

The cis isomer obtained from procedure described in Example 1-1 wasisolated by reverse phase HPLC (Sunfire C-18 column, 0.1% TFA inH₂O/CH₃CN) to provide(S)-2-[(R)-2-(3′-chloro-biphenyl-4-yl)-1-(1H-tetrazol-5-ylcarbamoyl)-ethylamino]-propionicacid ethyl ester; ¹H NMR (400 MHz, DMSO-d6) δ 1.07 (t, 3H, J=7.1 Hz),1.12 (d, 3H, J=6.8 Hz), 2.88 (dd, 1H, J=8.1, 13.6 Hz), 3.04 (dd, 1H,J=6.1, 13.6 Hz), 3.18-3.26 (m, 1H), 3.69-3.78 (m, 1H), 3.87-4.03 (m,2H), 7.35 (d, 2H, J=8.1 Hz), 7.37-7.42 (m, 1H), 7.47 (dd, 1H, J=7.8, 7.8Hz), 7.58-7.65 (m, 3H), 7.68-7.72 (m, 1H); MS: m/z (MH⁺) 443.

Following compounds were prepared using similar procedure as example 1-1and 2-1 with appropriate intermediates:

Ex- HPLC- am- RT MS ple (con- (M + # Product Intermediates dition) 1)Ex- am- ple 2-2

  (S)-2-[(R)-2-(2′,5′- Dichloro-biphenyl- 4-yl)-1-(1H-tetrazol-5-ylcarbamoyl)- ethylamino]- propionic acid tert- butyl ester

  (S)-2-((S)-1-tert- Butoxycarbonyl- ethylamino)-3-(2′,5′-dichloro-biphenyl- 4-yl)-propionic acid And  

  5-amino-1H- tetrazole 1.38 min (C) 505 Ex- am- ple 2-3

  (S)-2-[(R)-2-(3′-Chloro- biphenyl-4-yl)-1-(3- hydroxy-isoxazol-5-ylcarbamoyl)- ethylamino]-propionic acid ethyl ester

  (S)-2-((S)-1-tert- Butoxycarbonyl- ethylamino)-3-(3′-chloro-biphenyl-4- yl)-propionic acid And  

  5-amino-isoxazol-3-ol 1.46 min (C) 486 Ex- am- ple 2-4

  (S)-2-[(R)-2-(3′-Chloro- biphenyl-4-yl)-1-(1- methyl-1H-tetrazol-5-ylcarbamoyl)- ethylamino]-propionic acid ethyl ester

  (S)-3-(3′-Chloro- biphenyl-4-yl)-2-((S)- 1-ethoxycarbonyl-ethylamino)- propionic acid And  

  1-methyl-1H- tetrazol-5-ylamine 1.16 min (C) 457

Example 3-1(S)-2-[(S)-2-(3′-chloro-biphenyl-4-yl)-1-(1H-tetrazol-5-ylcarbamoyl)-ethylamino]-propionicacid

(S)-2-[(S)-2-(3′-chloro-biphenyl-4-yl)-1-(1H-tetrazol-5-ylcarbamoyl)-ethylamino]-propionicacid ethyl ester (100 mg, 0.226 mmol) was treated with 2M aqueous NaOH(2 mL) and EtOH (0.5 mL). After stirred at room temperature for 1 hour,the reaction mixture was acidified with 2M HCl to adjust pH 1. Theresulted precipitate was collected by filtration. The obtained materialwas crystallized from EtOH to give(S)-2-[(S)-2-(3′-chloro-biphenyl-4-yl)-1-(1H-tetrazol-5-ylcarbamoyl)-ethylamino]-propionicacid (94 mg).

¹H NMR (400 MHz, DMSO-d6) δ 1.15 (d, 3H, J=7.1 Hz), 2.94 (dd, 1H, J=7.3,13.7 Hz), 3.03 (dd, 1H, J=6.3, 13.6 Hz), 3.26 (dd, 1H, J=7.1, 13.9 Hz),3.81 (dd, 1H, J=6.9, 6.9 Hz), 7.33 (d, 2H, J=8.3 Hz), 7.38-7.42 (m, 1H),7.47 (dd, 1H, J=7.8, 7.8 Hz), 7.59-7.64 (m, 3H), 7.69 (dd, 1H, J=1.8,1.8 Hz), 15.9 (bs, 1H); MS: m/z (MH⁺) 415; HRMS: calculated forC₁₉H₁₉ClN₆O₃ (M)⁺414.1. found 414.1 Chiral HPLC retention time=13.17min. [condition: Daicel CHIRALPAK IA 4.6×100 mm); flow rate=1 ml/min.;eluent: 20% EtOH (with 0.1% TFA) in heptane].

Example 3-2(S)-2-[(S)-2-(2′,5′-dichloro-biphenyl-4-yl)-1-(1H-tetrazol-5-ylcarbamoyl)-ethylamino]-propionicacid

To a solution of(S)-2-[(S)-2-(2′,5′-dichloro-biphenyl-4-yl)-1-(1H-tetrazol-5-ylcarbamoyl)-ethylamino]-propionicacid tert-butyl ester (103 mg, 0.204 mmol) in DCM (2 mL) were added TFA(1 mL) and triethylsilane (0.098 mL, 0.611 mmol). After stirred for 8hours, the reaction mixture was concentrated under reduced pressure. Theresidue was purified by reverse phase HPLC (Sunfire C-18 column, eluent:0.1% TFA in H₂O/CH₃CN) to give(S)-2-[(S)-2-(2′,5′-dichloro-biphenyl-4-yl)-1-(1H-tetrazol-5-ylcarbamoyl)-ethylamino]-propionicacid. ¹H NMR (400 MHz, DMSO-d6+TFA-d) δ 1.49 (d, 3H, J=7.1 Hz), 3.29(dd, 1H, J=7.6, 13.9 Hz), 3.42 (dd, 1H, J=7.1, 14.2 Hz), 4.13 (dd, 1H,J=7.1, 14.0 Hz), 4.62 (dd, 1H, J=7.3, 7.3 Hz), 7.37 (d, 1H, J=2.5 Hz),7.37-7.43 (m, 2H), 7.40 (d, 2H, J=4.3 Hz), 7.48 (dd, 1H, J=2.5, 8.6 Hz),7.59 (d, 1H, J=8.6 Hz), 14.89 (bs, 1H); HPLC Retention time 1.25 minutes(condition A); MS: m/z (MH⁺) 449.

Following compounds were prepared using similar procedure as example 3-1or 3-2 with appropriate starting material and conditions:

Ex- HPLC- am- RT MS ple Hydrolysis (con- (M + # Product StartingMaterial Condition dition) 1) Ex- am- ple 3-3

  (S)-2-[(R)-2-(2′,5′- Dichloro-biphenyl- 4-yl)-1-(1H- tetrazol-5-ylcarbamoyl)- ethylamino]- propionic acid

  (S)-2-[(R)-2-(2′,5′- Dichloro- biphenyl-4-yl)-1-(1H-tetrazol-5-ylcarbamoyl)- ethylamino]- propionic acid tert-butyl esterTFA, triethyl- silane, DCM, RT 1.30 min. (A) 449 Ex- am- ple 3-4

  (S)-2-[(S)-2- Biphenyl-4-yl-1-(1H- tetrazol-5- ylcarbamoyl)-ethylamino]- propionic acid

  (S)-2-[(S)-2- Biphenyl-4-yl-1-(1H- tetrazol-5- ylcarbamoyl)-ethylamino]- propionic acid ethyl ester 1M LiOH, 2M NaOH aq, EtOH, RT1.28 min. (B) 381 Ex- am- ple 3-5

  (S)-2-[(S)-2-(5′- Fluoro-2-methoxy- biphenyl-4-yl)-1-(1H- tetrazol-5-ylcarbamoyl)- ethylamino]- propionic acid

  (S)-2-[(S)-2-(5′- Fluoro-2-methoxy- biphenyl-4-yl)-1-(1H- tetrazol-5-ylcarbamoyl)- ethylamino]- propionic acid tert-butyl ester TFA,triethyl- silane, DCM, RT 1.19 min. (A) 429 Ex- am- ple 3-6

  5-[(S)-3-Biphenyl-4- yl-2-((S)-1-carboxy- ethylamino)-propionylamino]-1H- pyrazole-3- carboxylic acid

  5-[(S)-3-Biphenyl-4- yl-2-((S)-1-tert- butoxycarbonyl- ethylamino)-propionylamino]-1H- pyrazole-3- carboxylic acid methyl ester 2M NaOH aq,EtOH RT. Followed by TFA, DCM, RT 1.26 min. (B) 423 Ex- am- ple 3-7

  (S)-2-[(S)-2-(3′- Chloro-biphenyl-4- yl)-1-(3-hydroxy- isoxazol-5-ylcarbamoyl)- ethylamino]- propionic acid

  (S)-2-[(S)-2-(3′- Chloro-biphenyl-4- yl)-1-(3-hydroxy- isoxazol-5-ylcarbamoyl)- ethylamino]- propionic acid tert-butyl ester TFA,triethyl- silane, DCM, RT 1.30 min. (A) 430 Ex- am- ple 3-8

  (S)-2-[(R)-2-(3′- Chloro-biphenyl-4- yl)-1-(3-hydroxy- isoxazol-5-ylcarbamoyl)- ethylamino]- propionic acid

  (S)-2-[(R)-2-(3′- Chloro-biphenyl-4- yl)-1-(3-hydroxy- isoxazol-5-ylcarbamoyl)- ethylamino]- propionic acid tert-butyl ester TFA,triethyl- silane, DCM, RT 1.40 min. (A) 430 Ex- am- ple 3-9

  (S)-2-{2-(3′-Chloro- biphenyl-4-yl)-1- [methyl-(1H-tetrazol-5-yl)-carbamoyl]- ethylamino}- propionic acid

  (S)-2-{2-(3′-Chloro- biphenyl-4-yl)-1- [methyl-(1H-tetrazol-5-yl)-carbamoyl]- ethylamino}- propionic acid ethyl ester 2M NaOH aq,EtOH, RT 1.16 min. (A) 429 Ex- am- ple 3-10

  (S)-2-[(S)-2-(3′- Chloro-biphenyl-4- yl)-1-(1-methyl-1H- tetrazol-5-ylcarbamoyl)- ethylamino]- propionic acid

  (S)-2-[(S)-2-(3′- Chloro-biphenyl-4- yl)-1-(1-methyl-1H- tetrazol-5-ylcarbamoyl)- ethylamino]- propionic acid ethyl ester 2M NaOH aq, EtOH,RT 1.38 min. (A) 429 Ex- am- ple 3-11

  (S)-2-[(R)-2-(3′- Chloro-biphenyl-4- yl)-1-(1-methyl-1H- tetrazol-5-ylcarbamoyl)- ethylamino]- propionic acid

  (S)-2-[(R)-2-(3′- Chloro-biphenyl-4- yl)-1-(1-methyl-1H- tetrazol-5-ylcarbamoyl)- ethylamino]- propionic acid ethyl ester 2M NaOH aq, EtOH,RT 1.43 min. (A) 429 Ex- am- ple 3-12

  (S)-2-[(S)-2-(3′- Chloro-biphenyl-4- yl)-1-(1H-tetrazol-5-ylcarbamoyl)- ethylamino]-4- phenyl-butyric acid

  (S)-2-[(S)-2-(3′- Chloro-biphenyl-4- yl)-1-(1H-tetrazol-5-ylcarbarnoyl)- ethylamino]-4- phenyl-butyric acid ethyl ester 2M NaOHaq, EtOH, RT 0.82 min. (C) 505 Ex- am- ple 3-13

  (S)-2-[(S)-2-(3′- Chloro-biphenyl-4- yl)-1-(1H-tetrazol-5-ylcarbamoyl)- ethylamino)-butyric acid

  (S)-2-[(S)-2-(3′- Chloro-biphenyl-4- yl)-1-(1H-tetrazol-5-ylcarbamoyl)- ethylamino]-butyric acid tert-butyl ester TFA, DCM, RT0.42 min. (C) 429 Ex- am- ple 3-14

  (S)-2-[(S)-2-(3′- Chloro-biphenyl-4- yl)-1-(1H-tetrazol-5-ylcarbamoyl)- ethylamino]-3- phenyl-propionic acid

  2-[2-(3′-Chloro- biphenyl-4-yl)-1- (1H-tetrazol-5- ylcarbamoyl)-ethylamino]- 3-phenyl- propionic acid ethyl ester 2M NaOH aq, EtOH, RT1.25 min. (A) 491 Ex- am- ple 3-15

  (S)-2-[(R)-2-(3′- Chloro-biphenyl-4- yl)-1-(1H-tetrazol-5-ylcarbamoyl)- ethylamino]-3- phenyl-propionic acid

  2-[2-(3′-Chloro- biphenyl-4-yl)-1- (1H-tetrazol-5- ylcarbamoyl)-ethylamino]-3-phenyl- propionic acid ethyl ester 2M NaOH aq, EtOH, RT1.28 min. (A) 491 Ex- am- ple 3-16

  (S)-3-benzyloxy-2- [(S)-2-(3′-chloro- biphenyl-4-yl)-1-(1H-tetrazol-5- ylcarbamoyl)- ethylamino]- propionic acid

  (S)-3-benzyloxy-2- [(S)-2-(3′-chloro- biphenyl-4-yl)-1-(1H-tetrazol-5- ylcarbamoyl)- ethylamino]- propionic acid ethyl ester 2MNaOH aq, EtOH, RT 1.35 min. (A) 521 Ex- am- ple 3-17

  (S)-2-[(S)-2-(3′- chloro-biphenyl-4- yl)-1-(1H-tetrazol-5-ylcarbamoyl)- ethylamino]-3- methoxy-propionic acid

  (S)-2-[(S)-2-(3′- chloro-biphenyl-4- yl)-1-(1H-tetrazol-5-ylcarbamoyl)- ethylamino]-3- methoxy-propionic acid ethyl ester 2M NaOHaq, EtOH, RT 0.93 min. (A) 445 Ex- am- ple 3-18

  (S)-2-[(S)-2-(3′- chloro-biphenyl-4- yl)-1-(1H-tetrazol-5-ylcarbamoyl)- ethoxy]-propionic acid

  (S)-2-[(S)-2-(3′- chloro-biphenyl-4- yl)-1-(1H-tetrazol-5-ylcarbamoyl)- ethoxy]-propionic acid ethyl ester 2M NaOH aq, MeOH, RT1.09 min (A) 416

Example 3-3

¹H NMR (400 MHz, DMSO-d6+TFA-d) δ 1.48 (d, 3H, J=7.1 Hz), 3.27 (dd, 1H,J=8.8, 13.1 Hz), 3.47 (dd, 1H, J=6.1, 13.4 Hz), 4.03 (dd, 1H, J=7.1,14.1 Hz), 4.47 (dd, 1H, J=7.3, 7.3 Hz), 7.37-7.42 (m, 5H), 7.47 (dd, 1H,J=2.8, 8.6 Hz), 7.58 (d, 1H, J=8.6 Hz), 14.89 (bs, 1H).

Example 3-4

¹H NMR (400 MHz, DMSO-d6) δ 1.37 (d, 3H, J=6.8 Hz), 3.20 (d, 2H, J=6.3Hz), 3.73-3.87 (bs, 1H), 4.25-4.38 (bs, 1H), 7.33-7.38 (m, 1H), 7.36 (d,2H, J=8.1 Hz), 7.45 (dd, 2H, J=7.4, 7.4 Hz), 7.60-7.66 (m, 4H).

Example 3-5

¹H NMR (400 MHz, DMSO-d6) δ 1.35-1.43 (m, 3H), 3.20 (bs, 2H), 3.71 (s,3H), 3.75-4.00 (m, 1H), 4.36 (bs, 1H), 7.05-7.20 (m, 3H), 7.31 (d, 2H,J=8.3 Hz), 7.45 (d, 2H, J=8.3 Hz).

Example 3-6

¹H NMR (400 MHz, DMSO-d6+TFA-d) δ 1.48 (d, 3H, J=7.3 Hz), 3.22 (dd, 1H,J=7.6, 13.6 Hz), 3.32 (dd, 1H, J=6.6, 13.9 Hz), 3.96 (dd, 1H, J=7.3,14.4 Hz), 4.50 (dd, 1H, J=7.3, 7.3 Hz), 6.97 (s, 1H), 7.33-7.38 (m, 3H),7.45 (t, 2H, J=7.8 Hz), 7.61-7.67 (m, 4H).

Example 3-7

¹H NMR (400 MHz, DMSO-d6) δ 1.37 (bd, 3H, J=4.8 Hz), 3.09-3.26 (m, 2H),3.67-3.90 (m, 1H), 4.10-4.37 (m, 1H), 5.83 (s, 1H), 7.34 (d, 2H, J=8.1Hz), 7.40-7.45 (m, 1H), 7.48 (dd, 1H, J=7.8, 7.8 Hz), 7.61-7.66 (m, 1H),7.66-7.73 (m, 3H).

Example 3-8

¹H NMR (400 MHz, DMSO-d6) δ 1.19-1.39 (m, 3H), 3.05-3.218 (m, 2H),3.30-4.25 (m, 2H), 5.83 (s, 1H), 7.33 (d, 2H, J=8.3 Hz), 7.40-7.44 (m,1H), 7.48 (dd, 1H, J=7.8, 7.8 Hz), 7.61-7.73 (m, 4H).

Example 3-9

¹H NMR (400 MHz, DMSO-d6) δ 1.48-1.57 (m, 3H), 3.05-3.47 (m, 2H),3.728/3.31 (s×2, total 3H), 4.02-4.21 (m, 1H), 5.61-5.82 (m, 1H),7.06-7.27 (m, 1H), 7.34-7.76 (m, 7H).

Example 3-10

¹H NMR (400 MHz, DMSO-d6) δ 1.35-1.43 (m, 3H), 3.13-3.34 (m, 2H),3.35-3.95 (m, 1H), 3.73 (s, 3H), 4.08-4.45 (m, 1H), 7.39-7.45 (m, 3H),7.49 (dd, 1H, J=7.8, 7.8 Hz), 7.62-7.75 (m, 4H).

Example 3-11

¹H NMR (400 MHz, DMSO-d6) δ 1.32-1.42 (m, 3H), 3.13-3.34 (m, 2H),3.35-3.95 (m, 1H), 3.73 (s, 3H), 4.02-4.36 (m, 1H), 7.37-7.45 (m, 3H),7.49 (dd, 1H, J=7.8, 7.8 Hz), 7.61-7.74 (m, 4H).

Example 3-12

¹H NMR (400 MHz, DMSO-d6) δ ppm 1.67-1.90 (m, 2H), 2.59 (t, J=7.7 Hz,2H), 2.96 (dd, J=13.6, 7.3 Hz, 1H), 3.07 (dd, J=13.6, 7.1 Hz, 1H),3.11-3.17 (m, 1H), 3.78 (t, J=7.1 Hz, 1H), 7.07-7.18 (m, 5H), 7.33 (d,J=8.3 Hz, 2H), 7.37-7.42 (m, 1H), 7.46 (t, J=8.0 Hz, 1H), 7.61 (d, J=8.3Hz, 3H), 7.68 (t, J=1.8 Hz, 1H), 12.02 (br. s., 1H), 15.89 (br. s., 1H).

Example 3-13

¹H NMR (400 MHz, DMSO-d₆) δ ppm 0.91 (t, J=7.5 Hz, 3H), 1.67-1.80 (m,2H), 3.08-3.27 (m, 2H), 3.56 (br. s., 3H), 4.16 (br. s., 1H), 7.34 (d,J=8.3 Hz, 2H), 7.41 (ddd, J=7.8, 2.0, 1.0 Hz, 1H), 7.47 (t, J=7.8 Hz,1H), 7.61 (dt, J=8.0, 1.5, 1.1 Hz, 1H), 7.64 (d, J=8.3 Hz, 2H), 7.68 (t,J=1.8 Hz, 1H), 12.27 (br. s., 1H), 16.09 (br. s., 1H).

Example 3-14

¹H NMR (400 MHz, DMSO-d₆) δ ppm 2.83-2.91 (m, 2H), 2.91-3.11 (m, 2H),3.56 (br. s., 2H), 3.88 (br. s., 1H), 7.14-7.20 (m, 3H), 7.20-7.26 (m,2H), 7.29 (d, J=8.3 Hz, 2H), 7.38-7.42 (m, 1H), 7.47 (t, J=7.8 Hz, 1H),7.56-7.63 (m, 3H), 7.68 (t, J=1.9 Hz, 1H), 11.92 (br. s., 1H), 15.91(br. s., 1H).

Example 3-15

¹H NMR (400 MHz, DMSO-d₆) δ ppm 2.81-3.03 (m, 4H), 3.51 (t, J=6.4 Hz,1H), 3.79 (t, J=6.2 Hz, 1H), 7.14-7.29 (m, 7H), 7.38-7.42 (m, 1H), 7.47(t, J=7.8 Hz, 1H), 7.55 (d, J=8.1 Hz, 2H), 7.59 (dt, J=7.6, 1.4 Hz, 1H),7.66 (t, J=1.9 Hz, 1H), 11.78 (br. s., 1H), 15.86 (br. s., 1H).

Example 3-16

¹H NMR (400 MHz, DMSO-d₆) δ ppm 2.97 (dd, 1H, J=7.1, 13.6 Hz), 3.07 (dd,1H, J=6.3, 13.6 Hz), 3.47 (dd, 1H, J=5.1, 5.1 Hz), 3.58 (d, 2H, J=5.1Hz), 3.87 (dd, 1H, J=6.6 Hz), 4.41 (d, 1H, J=12.4 Hz), 4.46 (d, 1H,J=12.1 Hz), 7.22-7.36 (m, 7H), 7.38-7.42 (m, 1H), 7.47 (t, 1H, j=7.8Hz), 7.58-7.64 (m, 3H), 7.68 (t, 1H, J=1.8 Hz).

Example 3-17

¹H NMR (400 MHz, DMSO-d₆) δ ppm 2.97-3.13 (m, 2H), 3.21 (s, 3H),3.40-3.61 (m, 3H), 3.76-4.01 (m, 1H), 7.34 (d, J=8.34 Hz, 2H), 7.38-7.43(m, 1H), 7.47 (t, J=8.08 Hz, 1H), 7.59-7.65 (m, 3H), 7.69 (bt, J=1.77Hz, 1H).

Example 3-18

¹H NMR (400 MHz, DMSO-d₆) d ppm 1.31 (d, J=6.6 Hz, 3H), 3.05-3.18 (m,2H), 4.03 (q, J=6.8 Hz, 1H), 4.58 (t, J=6.3 Hz, 1H), 7.35 (d, J=8.1 Hz,2H), 7.37-7.42 (m, 1H), 7.47 (t, J=7.8 Hz, 1H), 7.55-7.65 (m, 3H),7.66-7.72 (m, 1H), 12.13 (br. s., 1H), 12.69 (br. s., 1H), 15.96 (br.s., 1H)

Example 3-19

(S)-2-[(S)-2-(3′-chloro-biphenyl-4-O-1-(1H-tetrazol-5-ylcarbamoyl)-ethylamino]-3-hydroxy-propionicacid

Example 3-20

(S)-2-[(S)-2-biphenyl-4-yl-1-(1H-tetrazol-5-ylcarbamoyl)-ethylamino]-3-hydroxy-propionicacid

To a solution of(S)-3-benzyloxy-2-[(S)-2-(3′-chloro-biphenyl-4-yl)-1-(1H-tetrazol-5-ylcarbamoyl)-ethylamino]-propionicacid ethyl ester (47 mg, 0.090 mmol) in EtOAc (1 mL) and EtOH (1 mL) wasadded 5% Pd—C (9.6 mg, 0.0045 mmol). H₂ gas was loaded with a baloon andthe reaction mixture was stirred at 50° C. for 6 hours. The reactionmixture was filtered through celite pad and the filtrate wasconcentrated. The residue was purified by reverse phase HPLC (SunfireC-18 column, eluent: 0.1% TFA in H₂O/CH3CN) to give(S)-2-[(S)-2-(3′-chloro-biphenyl-4-yl)-1-(1H-tetrazol-5-ylcarbamoyl)-ethylamino]-3-hydroxy-propionicacid and(S)-2-[(S)-2-biphenyl-4-yl-1-(1H-tetrazol-5-ylcarbamoyl)-ethylamino]-3-hydroxy-propionicacid.

(S)-2-[(S)-2-(3′-chloro-biphenyl-4-yl)-1-(1H-tetrazol-5-ylcarbamoyl)-ethylamino]-3-hydroxy-propionicacid; NMR (400 MHz, DMSO-d6) δ ppm 2.99-3.14 (m, 2H), 3.50-3.67 (m, 3H),3.86-3.98 (m, 1H), 7.34 (d, 2H, J=8.3 Hz), 7.38-7.42 (m, 1H), 7.47 (t,2H, J=7.8 Hz), 7.58-7.70 (m, 4H)); HPLC Retention time 1.17 minutes(condition A); MS: m/z (MH+) 431.

(S)-2-[(S)-2-biphenyl-4-yl-1-(1H-tetrazol-5-ylcarbamoyl)-ethylamino]-3-hydroxy-propionicacid; NMR (400 MHz, DMSO-d6) δ ppm 3.18 (dd, 1H, J=7.6, 13.4 Hz),3.24-3.36 (m, 1H), 3.66-3.87 (m, 3H), 4.17-4.37 (m, 1H), 7.32 (d, 2H,J=8.1 Hz), 7.32-7.38 (m, 1H), 7.44 (t, 2H, J=7.8 Hz), 7.56-7.67 (m,4H)); HPLC Retention time 1.00 minutes (condition A); MS: m/z (MH+) 397.

Example 4-1(S)-3-(3′-chloro-biphenyl-4-yl)-2-((S)-2-methanesulfonylamino-1-methyl-2-oxo-ethylamino)-N-(1H-tetrazol-5-yl)-propionamide

Example 4-1 was prepared using similar procedure as example 1-1. NMR(400 MHz, DMSO-d6+TFA-d) δ 1.21 (d, J=6.32 Hz, 3H), 2.92-3.05 (m, 1H),3.05-3.14 (m, 1H), 3.17 (s, 3H), 3.34-3.46 (m, 1H), 3.82-3.95 (m, 1H),7.35 (d, J=8.08 Hz, 2H), 7.39-7.43 (m, 1H), 7.47 (t, J=7.83 Hz),7.60-7.66 (m, 3H), 7.68-7.22 (m, 1H)); HPLC Retention time 1.21 minutes(condition A); MS: m/z (MH+) 492.

Starting materials or intermediates were prepared in following manner:

Intermediate 1(S)-3-(3′-Chloro-biphenyl-4-yl)-2-((S)-1-ethoxycarbonyl-ethylamino)-propionicacid

Step 1: To a solution of Boc-L-4-bromophenylalanine (15.0 g, 43.6 mmol),3-chlorophenylboronic acid (8.52 g, 54.5 mmol), andtetrakis(triphenylphosphine)palladium(0) (1.51 g, 1.31 mmol) in1,2-dimethoxyethane (180 mL) was added 2M solution of aqueous NaCO₃ (33mL). The reaction mixture was heated to 85° C. After stirred for 2hours, the reaction mixture was cooled to room temperature and dilutedwith EtOAc. The mixture was washed with 1M HCl and brine. The organiclayer was dried over Na₂SO₄, concentrated under reduced pressure, andpurified by silica gel column chromatography (eluent: 10% MeOH indichloromethane) to give(S)-2-tert-butoxycarbonylamino-3-(3′-chloro-biphenyl-4-yl)-propionicacid (13.6 g). ¹H NMR (400 MHz, CDCl₃) δ 1.43 (s, 9H), 3.08-3.17 (m,1H), 3.21-3.31 (m, 1H), 4.65 (bs, 1H), 5.01 (bs, 1H), 7.23-7.32 (m, 3H),7.45-7.50 (m, 2H), 7.52-7.60 (m, 1H), 7.63-7.70 (m, 2H); MS: m/z (MW)376.

Step 2: To a solution of(S)-2-tert-butoxycarbonylamino-3-(3′-chloro-biphenyl-4-yl)-propionicacid (12.9 g, 34.3 mmol) in DMF (130 mL) were added benzyl bromide (8.16mL, 68.6 mmol) and NaHCO₃ (5.77 g, 68.6 mmol). After stirred at roomtemperature overnight, the reaction mixture was diluted with EtOAc. Themixture was washed with H₂O and brine, dried over Na₂SO₄, andconcentrated under reduced pressure. The obtained residue was treatedwith 4M HCl in dioxane (30 mL) and stirred for 2 hours. The reactionmixture was concentrated and the resulted residue was rinsed with iPr₂Oto give (S)-2-amino-3-(3′-chloro-biphenyl-4-yl)-propionic acid benzylester (11.2 g). ¹H NMR (400 MHz, DMSO-d6) δ 3.14 (dd, 1H, J=7.7, 12.0Hz), 3.27 (dd, 1H, J=5.9, 12.0 Hz), 4.38 (dd, 1H, J=5.9, 7.7 Hz), 5.15(s, 2H), 7.23-7.27 (m, 2H), 7.30-7.34 (m, 5H), 7.42-7.45 (m, 1H), 7.51(dd, 1H, J=7.6, 7.6 Hz), 7.61-7.66 (m, 3H), 7.69 (dd, 1H, J=1.8, 1.8Hz), 8.64 (bs, 2H); MS: m/z (MW) 366.

Step 3: To a solution of(S)-2-amino-3-(3′-chloro-biphenyl-4-yl)-propionic acid benzyl ester(10.0 g, 24.9 mmol) in dichloromethane (100 mL) was added triethylamine(10.4 mL, 74.6 mmol) at 0° C. After stirred for 10 min, ethyl(R)-2-(trifluoromethylsulfonyloxy)propionate (9.3 mL, 49.5 mmol) wasadded at room temperature and stirred for 1 hour. Additionaltriethylamine (10.4 mL, 74.6 mmol) and ethyl(R)-2-(trifluoromethylsulfonyloxy)propionate (9.3 mL, 49.5 mmol) wereadded at room temperature and stirred for additional 2 hours. Thereaction mixture was washed with H₂O and the organic layer wasconcentrated under reduced pressure. The obtained residue was purifiedby silica gel column chromatography (EtOAc/heptane) to give(S)-3-(3′-chloro-biphenyl-4-yl)-2-((S)-1-ethoxycarbonyl-ethylamino)-propionicacid benzyl ester (10.6 g). ¹H NMR (400 MHz, CDCl₃) δ 1.21 (t, 3H, J=7.3Hz), 1.27 (d, 3H, J=6.8 Hz), 1.89 (bs, 1H), 2.95-3.07 (m, 2H), 3.38 (dd,1H, J=6.8, 14.8 Hz), 3.69 (dd, 1H, J=7.1, 7.1 Hz), 4.06-4.17 (m, 2H),5.06 (d, 1H, J=12.1 Hz), 5.12 (d, 1H, J=12.1 Hz), 7.20-7.25 (m, 4H),7.28-7.34 (m, 4H), 7.35 (dd, 1H, J=7.6, 7.6 Hz), 7.41-7.46 (m, 3H), 7.53(dd, 1H, J=1.5, 1.5 Hz); MS: m/z (MW) 466.

Step 4: A suspension of(S)-3-(3′-chloro-biphenyl-4-yl)-2-(S)-1-ethoxycarbonyl-ethylamino)-propionicacid benzyl ester (10.0 g, 21.5 mmol) and 5% Pd on carbon (0.914 g) inEtOAc (200 mL) was treated with H₂ (balloon) and stirred at 10-15° C.for 1.5 hour and at room temperature for 0.5 hour. The resultedprecipitate was dissolved in methanol and filtered through celite pad.The filtrate was concentrated under reduced pressure and the obtainedresidue was re-crystallized from EtOAc to give(S)-3-(3′-chloro-biphenyl-4-yl)-2-(S)-1-ethoxycarbonyl-ethylamino)-propionicacid (5.6 g). The mother liquor was concentrated under reduced pressureand purified by silica gel column chromatography to give additionalamount of(S)-3-(3′-chloro-biphenyl-4-yl)-2-(S)-1-ethoxycarbonyl-ethylamino)-propionicacid (1.4 g). ¹H NMR (400 MHz, DMSO-d6) δ 1.13 (t, 3H, J=7.1 Hz), 1.15(d, 3H, J=6.8 Hz), 2.85 (dd, 1H, J=7.1, 14.1 Hz), 2.93 (dd, 1H, J=6.3,13.6 Hz), 3.30-3.37 (m, 1H), 3.48 (dd, 1H, J=6.5, 6.5 Hz), 4.03 (q, 2H,J=7.1 Hz), 7.32 (d, 2H, J=8.3 Hz), 7.38-7.43 (m, 1H), 7.48 (dd, 1H,J=7.8, 7.8 Hz), 7.59-7.65 (m, 3H), 7.70 (dd, 1H, J=2.0, 2.0 Hz); MS: m/z(MH⁺) 376.

Intermediate 2(S)-2-((S)-1-tert-Butoxycarbonyl-ethylamino)-3-(2′,5′-dichloro-biphenyl-4-yl)-propionicacid

Same procedures described in step 1 (2,5-dichlorophenylboronic acid wasused instead of 3-chlorophenylboronic acid) and step 2 for thepreparation of intermediate 1 were used to prepare((S)-2-amino-3-(2′,5′-dichloro-biphenyl-4-yl)-propionic acid benzylester hydrochloride.

Step 3′: t-Butyl (R)-2-(trifluoromethylsulfonyloxy)propionate wasprepared from (R)-2-hydroxy-propionic acid tert-butyl ester (602 mg,4.12 mmol), triflic anhydride (0.696 mL, 4.12 mmol) and 2,6-lutidine(0.480 mL, 4.12 mmol) in DCM (5 mL). To a suspension of((S)-2-amino-3-(2′,5′-dichloro-biphenyl-4-yl)-propionic acid benzylester hydrochloride (600 mg, 1.38 mmol) in dichloromethane (10 mL) wasadded triethylamine (0.574 mL, 4.12 mmol) at 0° C. After stirred for 10min, a half amount of the freshly prepared t-butyl(R)-2-(trifluoromethylsulfonyloxy)propionate was added at roomtemperature and stirred for 1 hour. Additional triethylamine (0.574 mL,4.12 mmol) and the rest of t-butyl(R)-2-(trifluoromethylsulfonyloxy)propionate were added at roomtemperature and stirred for additional 2 hours. The reaction mixture waswashed with H₂O and the organic layer was concentrated under reducedpressure. The obtained residue was purified by silica gel columnchromatography (EtOAc/heptane) to give(S)-2-(S)-1-tert-butoxycarbonyl-ethylamino)-3-(2′,5′-dichloro-biphenyl-4-yl)-propionicacid benzyl ester (580 mg). ¹H NMR (400 MHz, CDCl₃) δ 1.24 (t, 3H, J=6.8Hz), 1.41 (s, 9H), 3.00-3.07 (m, 2H), 3.26 (dd, 1H, J=7.1, 13.9 Hz),3.70 (dd, 1H, J=7.1, 7.1 Hz), 5.09 (s, 2H), 7.20-7.42 (m, 12H); MS: m/z(MH⁺) 528.

Step 4′: A suspension of(S)-2-(S)-1-tert-Butoxycarbonyl-ethylamino)-3-(2′,5′-dichloro-biphenyl-4-yl)-propionicacid benzyl ester (580 mg, 1.10 mmol) and 5% Pd on carbon (0.146 g) inEtOAc (10 mL) was treated with H₂ (balloon) and stirred at rt for 1.5hour. The resulted precipitate was dissolved in methanol and filteredthrough celite pad. The filtrate was concentrated under reduced pressureand the obtained residue was re-crystallized from EtOAc to give(S)-2-((S)-1-tert-butoxycarbonyl-ethylamino)-3-(2′,5′-dichloro-biphenyl-4-yl)-propionicacid (438 mg). ¹H NMR (400 MHz, DMSO-d6) δ 1.12 (d, 3H, J=7.1 Hz), 1.35(s, 9H), 2.84 (dd, 2H, J=7.3, 13.6 Hz), 2.95 (dd, 2H, J=6.1, 13.6 Hz),3.20 (dd, 1H, J=6.8, 13.6 Hz), 3.48 (dd, 1H, J=6.1, 7.3 Hz), 7.33 (d,2H, J=8.6 Hz), 7.37 (d, 2H, J=8.3 Hz), 7.42-7.49 (m, 2H), 7.60 (d, 2H,J=8.6 Hz); MS: m/z (MH⁺) 438.

Following intermediates were prepared using similar procedure asintermediate 1 or intermediate 2 with appropriate reagent:

HPLC-RT MS Intermediate # Intermediate Reagent (condition) (M + 1)Intermediate 3

  (S)-3-Biphenyl-4-yl-2- ((S)-1-ethoxy carbonyl-ethylamino)- propionicacid phenylboronic acid was used instead of 3- chlorophenylboronic acidin step 1 0.71 min (C) 342 Intermediate 4

  (S)-2-((S)-1-tert- Butoxycarbonyl- ethylamino)-3-(5′-fluoro-2′-methoxy- biphenyl-4-yl)-propionic acid 2-methoxy-5-fluoro-phenylboronic acid was used instead of 3- chlorophenylboronic acid instep 1 1.07 min (C) 418 Intermediate 5

  (S)-3-Biphenyl-4-yl-2- ((S)-1-tert- butoxycarbonyl-ethylamino)-propionic acid phenylboronic acid was used instead of 3-chlorophenylboronic acid in step 1 1.05 min (C) 370 Intermediate 6

  (S)-2-[(S)-1-carboxy- 2-(3′-chloro-biphenyl- 4-yl)-ethylamino]-4-phenyl-butyric acid ethyl ester (R)-2-Hydroxy-4-phenyl- butyric acidethyl ester was used instead of (R)- 2-hydroxy-propionic acid tert-butylester in Step 3′ 1.39 min (C) 466 Intermediate 7

  (S)-2-[(S)-1-Carboxy- 2-(3′-chloro-biphenyl- 4-yl)-ethylamino]-butyric acid tert-butyl ester (R)-2-Hydroxy-butyric acid tert-butylester was used instead of (R)-2- hydroxy-propionic acid tert-butyl esterin Step 3′ 1.15 min (C) 418 Intermediate 8

  (S)-3-(3′-Chloro- biphenyl-4-yl)-2- ((S)-1-ethoxycarbonyl-2-phenyl-ethyl amino)-propionic acid (R)-2-Hydroxy-3-phenyl- propionicacid ethyl ester was used instead of (R)- 2-hydroxy-propionic acidtert-butyl ester in Step 3′ 1.27 min (C) 452

Intermediate 9 [1-(4-Methoxy-benzyl)-1H-tetrazol-5-yl]-methyl-amine

To a suspension of 5-amino-1H-tetrazole (1.50 g, 17.6 mmol) in DMF (30mL) were added Cs₂CO₃ (8.62 g, 26.4 mmol) and PMBCl (2.90 g, 18.5 mmol).After stirred at 60° C. for 3 hours, the reaction mixture was cooled toroom temperature and diluted with EtOAc. The mixture was washed with H₂Oand brine, dried over Na₂SO₄, and concentrated under reduced pressure.The residue was diluted with DCM and the resulted precipitate wascollected by filtration to give1-(4-methoxy-benzyl)-1H-tetrazol-5-ylamine (0.625 g). ¹H NMR (400 MHz,DMSO-d6) δ 3.73 (s, 3H), 5.27 (s, 2H), 6.78 (s, 2H), 6.92 (d, 2H, J=8.8Hz), 7.21 (d, 2H, J=8.8 Hz).

Next, to a suspension of 1-(4-methoxy-benzyl)-1H-tetrazol-5-ylamine (600mg, 2.92 mmol) in MeOH (10 mL) were added paraformaldehyde (132 mg, 4.39mmol) and sodium methoxide (632 mg, 25 wt % in MeOH). The mixture wasrefluxed for 30 min until the suspension turned into a clear solution.The mixture was cooled to room temperature and sodium borohydride (332mg, 8.77 mmol) was added portionwise. The reaction mixture was refluxedagain for 15 min. After cooled to room temperature, the reaction wasquenched with H₂O. The mixture was diluted with EtOAc, partiallyconcentrated, and washed with brine. The organic layer was dried overNa₂SO₄ and concentrated under reduced pressure. The residue was purifiedby silica gel column chromatography (eluent: 10% MeOH in DCM) to give[1-(4-methoxy-benzyl)-1H-tetrazol-5-yl]-methyl-amine (0.63 g). ¹H NMR(400 MHz, CDCl₃) δ 3.00 (d, 3H, J=5.3 Hz), 3.61 (bs, 1H), 3.82 (s, 3H),5.25 (s, 2H), 6.91 (d, 2H, J=8.8 Hz), 7.16 (d, 2H, J=8.8 Hz); MS: m/z(MH⁺) 220.

Following intermediates were prepared using similar procedure asintermediate 1 or intermediate 2 with appropriate reagent:

HPLC-RT MS Intermediate # Intermediate Reagent (condition) (M + 1)Intermediate 9

  (S)-2-((S)-2- benzyloxy-1- ethoxycarbonyl- ethylamino)-3-(3′-chloro-biphenyl-4-yl)- propionic acid (R)-3-Benzyloxy-2-hydroxy-propionic acid ethyl ester was used instead of (R)-2-hydroxy-propionic acid ethyl ester in Step 3 1.41 min (C) 482 Intermediate 10

  (S)-3-(3′-chloro- biphenyl-4-yl)-2-((S)- 1-ethoxycarbonyl-2-methoxy-ethylamino)- propionic acid (R)-2-Hydroxy-3- methoxy-propionicacid ethyl ester was used instead of (R)-2-hydroxy- propionic acid ethylester in Step 3 0.56 min (C) 496

Intermediate 11(S)-3-(3′-chloro-biphenyl-4-yl)-2-((S)-1-ethoxycarbonyl-ethoxy)-propionicacid

Step 1: To a mixture of 4-bromo-L-phenylalanine (2.5 g, 10.24 mmol) andthe solvent of acetic acid (20 ml) and water (75 ml) in an ice bath wasadded dropwise a solution of sodium nitrite (2.120 g, 30.7 mmol) inwater (20.00 ml). The mixture was slowly warmed up to room temperatureand stirred overnight. To the suspension was added methylamine in THF(20.48 ml, 41.0 mmol) dropwise slowly and the mixture turned to clearand stirred at room temperature for 1 hr. The mixture was concentratedto remove THF and extracted with EtOAc. The combined organic layer waswashed with brine and dried over anhydrous sodium sulfate, filtered andconcentrated to give the crude as off white solid: 1.7 g (yield: 43%).HPLC retention time=0.83 minutes (condition A); MS (m+2)=246.

Step 2: To a solution of (S)-3-(4-bromo-phenyl)-2-hydroxy-propionic acid(1.5 g, 6.12 mmol) in DME (60 ml) at room temperature was added3-chlorobenzeneboronic acid (1.436 g, 9.18 mmol) and followed by aq.Na₂CO₃ (6.12 ml, 12.24 mmol) and Pd(Ph₃P)₄ (0.212 g, 0.184 mmol). Themixture was stirred at 85° C. overnight. The reaction was added moreEtOAc and acidified by 1N HCl to PH˜5. The combined organic layer waswashed with brine and dried over anhydrous sodium sulfate, filtered andconcentrated. The residue was purified by HPLC (20 to 80% ACN—H₂O (0.1%TFA)) to give the white solid: 550 mg (yield: 32%). HPLC retentiontime=1.23 minutes (condition A); MS (m⁻¹)=275.

Step 3: To a solution of(S)-3-(3′-chloro-biphenyl-4-yl)-2-hydroxy-propionic acid benzyl ester(282 mg, 0.769 mmol) in THF (6 ml) at −78° C. was added LiHMDS/THF(1.999 ml, 1.999 mmol) and the resulting yellow mixture was stirred at−78° C. for 25 mins then was added (R)-ethyl2-(trifluoromethylsulfonyloxy)propanoate (0.860 ml, 4.61 mmol) at −20°C. 1 hr the reaction was almost complete. The reaction was quenched bysat. NH4Cl and was extracted with EtOAc. The combined organic layer waswashed with brine, filtered and concentrated. The residue was purifiedby HPLC (75 to 100% ACN—H₂O (0.1% TFA)) to give the product: 140 mg(yield: 39%). HPLC retention time=1.57 minutes (condition C); MS(m+1)=467.

Step 4: A mixture of(S)-3-(3′-chloro-biphenyl-4-yl)-2-((S)-1-ethoxycarbonyl-ethoxy)-propionicacid benzyl ester and 10% Pd/C wet in EtOAc was hydrogenated under H₂baloon for 30 mins. The reaction was filtered off the catalyst andconcentrated. The residue was purified by HPLC (15 to 70% ACN—H₂O (0.1%TFA)) to give oil: 128 mg. HPLC retention time=1.07 minutes (conditionC); MS (m−1)=375.

Intermediate 12(S)-3-(3′-chloro-biphenyl-4-yl)-2-((S)-2-methanesulfonylamino-1-methyl-2-oxo-ethylamino)-propionicacid

Step 1: To a solution of(S)-2-(S)-1-tert-butoxycarbonyl-ethylamino)-3-(3′-chloro-biphenyl-4-yl)-propionicacid benzyl ester (1.12 g, 2.27 mmol) in DCM (5 mL) was added TFA (5mL). After being stirred for 3 hours, the reaction mixture wasconcentrated and purified by silica gel column chromatography (eluent:10% MeOH in DCM) to give(S)-2-((S)-1-carboxy-ethylamino)-3-(3′-chloro-biphenyl-4-yl)-propionicacid benzyl ester. MS: m/z (MH⁺) 438; HPLC retention time 0.73 min (HPLCcondition C).

Step 2: To a solution of(S)-2-((S)-1-carboxy-ethylamino)-3-(3′-chloro-biphenyl-4-yl)-propionicacid benzyl ester (600 mg, 1.37 mmol) in DCM (7 mL) and saturatedaqueous NaHCO₃ solution (2 mL) was added triphosgene (407 mg, 1.37mmol). After being stirred for 0.5 hours, the reaction mixture wasdiluted with EtOAc and stirred for additional 0.5 hours until generationof gas was completed. The organic layer was separated, washed with brineand concentrated. This was dissolved in DCM (7 mL) andmethanesulfonamide (195 mg, 2.06 mmol) was added. After being stirred atrt for 1 hour, the reaction mixture was diluted with EtOAc and washedwith brine. The organic layer was dried over Na₂SO₄, concentrated andpurified by silica gel column chromatography (eluent: 10% MeOH in DCM)to give(S)-3-(3′-chloro-biphenyl-4-yl)-2-((S)-2-methanesulfonylamino-1-methyl-2-oxo-ethylamino)-propionicacid benzyl ester. MS: m/z (MW) 515; HPLC retention time 1.58 min (HPLCcondition A).

Step 3: This was dissolved in EtOAc. 5% Pd—C (146 mg) was added andhydrogenated with H₂ balloon at rt for 1 hour. The reaction mixture wasfiltered through celite pad and the filtrate was concentrated. Theresultant solid was re-crystallized from MeOH to give(S)-3-(3′-chloro-biphenyl-4-yl)-2-((S)-2-methanesulfonylamino-1-methyl-2-oxo-ethylamino)-propionicacid. MS: m/z (MH⁺) 425; HPLC retention time 1.14 min (HPLC conditionA).

Intermediate 12-2(S)-2-((S)-1-tert-butoxycarbonyl-ethylamino)-3-(3′-chloro-biphenyl-4-yl)-propionicacid benzyl ester

Intermediate 12-2 was prepared using similar procedure as intermediate 1and intermediate 2 with appropriate reagent. MS: m/z (MH⁺) 494; HPLCretention time 1.50 min (HPLC condition C).

It can be seen that the compounds of the invention are useful asinhibitors of Neutral endopeptidase (EC 3.4.24.11) activity andtherefore useful in the treatment of diseases and conditions associatedwith Neutral endopeptidase (EC 3.4.24.11) activity such as the diseasesdisclosed herein.

It will be understood that the invention has been described by way ofexample only and modifications may be made whilst remaining within thescope and spirit of the invention.

What is claimed is:
 1. A method of treating a disorder or a diseaseassociated with neutral endopeptidase EC. 3.4. 24.11. activity in asubject in need thereof, comprising: administering to the subject atherapeutically effective amount of a compound of the formula (I′):

wherein: X¹ is OH, —O—C₁₋₇alkyl, —NR^(a)R^(b), —NHS(O)₂—C₁₋₇alkyl or—NHS(O)₂-benzyl, wherein R^(a) and R^(b) for each occurrence areindependently H or C₁₋₇alkyl; R¹ is H, C₁₋₆ alkyl or C₆₋₁₀-aryl-C₁₋₆alkyl, wherein alkyl is optionally substituted with benzyloxy, hydroxyor C₁₋₆ alkoxy; for each occurrence, R² is independently C₁₋₆-alkoxy,hydroxy, halo, C₁₋₆-alkyl, cyano or trifluoromethyl; A² is O or NR⁵; R⁴and R⁵ are independently H or C₁₋₆ alkyl; A¹ is a bond or C₁₋₃alkylenechain; R³ is a 5- or 6-membered heteroaryl, C₆₋₁₀-aryl orC₃₋₇-cycloalkyl, wherein each heteroaryl, aryl or cycloalkyl areoptionally substituted with one or more groups independently selectedfrom the group consisting of C₁₋₆alkyl, halo, haloC₁₋₆alkyl, C₁₋₆alkoxy,hydroxy, CO₂H and CO₂C₁₋₆alkyl; R⁶ for each occurrence is independentlyhalo, hydroxy, C₇₋₇alkoxy, halo, C₁₋₇alkyl or halo-C₁₋₇alkyl; or R⁴,A¹-R³, together with the nitrogen to which R⁴ and A¹-R³ are attached,form a 4- to 7-membered heterocyclyl or a 5- to 6-membered heteroaryl,each of which is optionally substituted with one or more groupsindependently selected from the group consisting of C₁₋₆alkyl, halo,haloC₁₋₆alkyl, C₁₋₆alkoxy, hydroxy, CO₂H and CO₂C₁₋₆alkyl; and m is 0 oran integer from 1 to 5; s is 0 or an integer from 1 to 4; or apharmaceutically acceptable salt thereof, wherein the disorder ordisease is selected from, left ventricular hypertrophy, angina, renalinsufficiency, renal failure, diabetic nephropathy, non-diabeticnephropathy, nephroic syndrome, glomerulonephritis, scleroderma,glomerular sclerosis, protein urea of primary renal disease, renalvascular hypertention, diabetic retinopathy and end-stage renal disease(ESRD), endothelial dysfunction, diastolic dysfunction, hypertrophiccardiomyopathy, diabetic cardiac myopathy, supraventricular andventricular arrhythmias, atrial fibrillation (AF), cardiac fibrosis,atrial flutter, detrimental vascular remodeling, plaque stabilization,myocardial infarction (MI), renal fibrosis, polycystic kidney disease(PKD), renal failure, cyclical oedema, Menières disease,hyperaldosteroneism hypercalciuria, ascites, glaucoma, menstrualdisorders, preterm labour, pre-eclampsia, endometriosis, andreproductive disorders, asthma, obstructive sleep apnea, inflammation,leukemia, pain, epilepsy, affective disorders, depression, psychoticcondition, dementia, geriatric confusion, obesity, gastrointestinaldisorders, wound healing, septic shock, gastric acid secretiondysfunction, hyperreninaemia, cystic fibrosis, restenosis, type-2diabetes, metabolic syndrome, diabetic complications, atherosclerosis,male and female sexual dysfunction.
 2. The method according to claim 1wherein the compound is of Formula I:

wherein: X¹ represent OH or O—C₁₋₆-alkyl; R¹ is H, C₁₋₆ alkyl orC₆₋₁₀-aryl-C₁₋₆ alkyl; for each occurrence, R² is independentlyC₁₋₆-alkoxy, hydroxy, halo, C₁₋₆-alkyl, cyano or trifluoromethyl; R⁴ andR⁵ are independently H or C₁₋₆ alkyl; A¹ is a bond or C₁₋₃alkylenechain; R³ is a 5- or 6-membered heteroaryl, C₆₋₁₀-aryl orC₃₋₇-cycloalkyl, wherein each heteroaryl, aryl or cycloalkyl areoptionally substituted with one or more groups independently selectedfrom the group consisting of C₇₋₆alkyl, halo, haloC₁₋₆alkyl, C₁₋₆alkoxy,hydroxy, CO₂H and CO₂C₁₋₆alkyl; R⁶ for each occurrence is independentlyhalo, hydroxy, C₁₋₇alkoxy, halo, C₁₋₇alkyl or halo-C₁₋₇alkyl; or R⁴,A¹-R³, together with the nitrogen to which R⁴ and A¹-R³ are attached,form a 4- to 7-membered heterocyclyl or a 5- to 6-membered heteroaryl,each of which is optionally substituted with one or more groupsindependently selected from the group consisting of C₁₋₆alkyl, halo,haloC₇₋₆alkyl, C₁₋₆alkoxy, hydroxy, CO₂H and CO₂C₁₋₆alkyl; and m is 0 oran integer from 1 to 5; s is 0 or an integer from 1 to 4; or apharmaceutically acceptable salt thereof.
 3. The method of claim 1wherein the compound is of Formula II:

or a pharmaceutically acceptable salt thereof.
 4. The method of claim 1wherein the compound is of Formula III:

Wherein X² is halo and p is 0 or an integer from 1 to 4, or apharmaceutically acceptable salt thereof.
 5. The method according toclaim 4 wherein the compound is of formula IV:

or a pharmaceutically acceptable salt thereof.
 6. The method accordingto claim 1 wherein X¹ represent OH or O—C₁₋₆-alkyl; R¹ is H or C₁₋₆alkyl; R² for each occurrence is independently C₁₋₆-alkoxy, hydroxy,halo, C₁₋₆-alkyl, cyano or trifluoromethyl; R⁴ and R⁵ are independentlyH or C₁₋₆ alkyl; A¹ is a bond or C₁₋₃alkylene chain; R³ is a 5- or6-membered heteroaryl optionally substituted with one or moresubstituents independently selected from the group consisting ofC₁₋₆alkyl, halo, haloC₁₋₆alkyl, C₁₋₆alkoxy, hydroxy, CO₂H andCO₂C₁₋₆alkyl; R⁶ for each occurrence is independently halo, hydroxy,C₇₋₇alkoxy, halo, C₁₋₇alkyl or halo-C₁₋₇alkyl; m is 0 or an integer of 1to 5; s is 0 or an integer of 1 to 4; or a pharmaceutically acceptablesalt thereof.
 7. The method of claim 1 wherein A¹ is a bond or CH₂; or apharmaceutically acceptable salt thereof.
 8. The method of claim 1wherein R¹ is methyl or ethyl, R⁵ and R⁴ are H; or a pharmaceuticallyacceptable salt thereof.
 9. The method of claim 1 wherein R³ is a5-membered ring heteroaryl selected from the group consisting ofoxazole, pyrrole, pyrazole, isooxazole, triazole, tetrazole, oxadiazole,thiazole, isothiazole, thiophene, imidazole and thiadiazole; each ofwhich is optionally substituted with one or more substituentsindependently selected from the group consisting of C₁₋₆alkyl, halo,haloC₁₋₆alkyl, C₁₋₆alkoxy, hydroxy, CO₂H and CO₂C₁₋₆alkyl; or apharmaceutically acceptable salt thereof.
 10. The method of claim 9wherein R³ is tetrazole; or a pharmaceutically acceptable salt thereof.11. The method of claim 5 wherein X² is Cl, or a pharmaceuticallyacceptable salt thereof. 12-19. (canceled)